Conductive yarn

ABSTRACT

An electrically conductive yarn or film and method of manufacturing thereof in which a SP1/nanoparticle complex bound to the yarn or film serves as a platform for adhesion of a metallic coating.

CROSS-REFERENCE TO RELATED APPLICATIONS

This application is a National Phase Application of PCT InternationalApplication No. PCT/IL2017/050106, International Filing Date Jan. 29,2017, claiming priority from IL Patent Application(s) No(s). 243839,filed Jan. 28, 2016, which are hereby incorporated by reference in theirentirely.

BACKGROUND OF THE INVENTION

The present invention relates to the enablement of non-metallic yarns,fabrics and films as electrical conductors by supporting a coating ofconductive metal with a Stable Protein (SP1) nanoparticle matrix boundto the fabric or film material.

Stable Protein 1 (SP1)

Stable protein 1 (SP1) is a homo-oligomeric protein isolated from aspen(Populus tremula aspen) plants and forms a ring-shape dodecamericparticle with a central cavity. The oligomeric form of SP1 is anexceptionally stable structure that is resistant to proteases, such astrypsin, V8, and proteinase K, high temperatures, organic solvents, andhigh levels of ionic detergent.

Carbon Nanoparticles (CNP) Reinforced Composite Materials

Although the present invention relates both to carbon and non-carbonnanoparticles, without diminishing in scope carbon nanoparticles will bediscussed herewithin.

Carbon nanotubes are nano-scale hollow cylinders of graphite carbonatoms. They provide the highest Young's modulus (stiffness), highestthermal conductivity, highest electrical conductivity, and highestcurrent density of any known material, while having a low density. Thenanotubes may consist of one (single walled carbon nanotubes) up to tens(multi-walled CNTs) and hundreds of concentric shells of carbons withadjacent shells separation of ˜0.34 nm. Single walled carbon nanotubestend to be stronger, more flexible, more transparent and betterelectrical conductors and are more transparent. High production costsplus health and safety considerations have led to multi-walled carbonnanotubes being more widely used in composite materials.

When carbon nanotubes are added to a matrix material the compositenormally takes on some of the carbon nanotubes' properties, due to therule of mixtures. However, the theoretical property values of carbonnanotubes composites are presently not attained due to the inability toefficiently produce fully integrated composites.

SP1 variants capable of forming molecular complexes with carbonnanotubes address the insufficient bonding across the interface of thenanotube and matrix material.

Carbon Nanoparticles Binding with Fabrics

Carbon Nanoparticles possess much larger surface specific area thanfibers, fabrics, or films, and is therefore a desirable platform forloading conductive metals. SP1 protein facilitates binding of CNP withthe fibers, even when low loading levels of SP1 are employed. SP1binding to the fiber has been found to be enhanced through a priorapplication of a polymeric coating on the fiber via bonding of reactivegroups of the protein.

Specifically designed SP1/CNP compositions (Also referred to as matricesand complexes) utilizing carbon black (CB), carbon nanotubes (CNT) ofeither graphene or graphite form stable dispersions in solvents. Theymay be complexed to a broad range of target compounds such as carbonfabrics, aramid, polyester, or nylon fibers, yarns, films, fabrics, andalso glass fiber fabrics to form useful molecular complexes in theproduction of highly specific composite materials such asSP1-polypeptide-CNP-aramid complex fabrics, films, yarns and polymericfabrics.

Carbon Nanoparticles Binding with Aramid Polymers (Kevlar)

As is well-known in the art, aramid (e.g. KEVLAR™) is chemically inertand is not soluble in any common solvent, has a very high melting point,and decomposes above 400° C. As a result, aramid fibers must be producedby wet spinning from sulfuric acid solutions. Binding of SP1/CNP complexto aramid can facilities the adhesion of carbon nanoparticles (e.g., CNTor CB) onto pre-formed polymer products like KEVLAR™ yarns.

Electrical Conductivity

The SP1/CNP complex further provides a platform for bonding electricallyconductive metals to yarn or fabric. To date, conductive fabrics havebeen constructed by spinning metallic yarn together with staple yarn or,in the production of conductive fabric, incorporating metallic wire ormesh into fabric. The SP1/CNP platform advantageously facilitiesscalable processing schemes and highly efficient use of metallicmaterials by exhausting the metallic solutions from their metalliccontent during conductive coating processes. The SP1/CNP platformfurther provides flexibility and shape memory functionality.

SUMMARY OF THE INVENTION

According to the teachings of the present invention there is provided aconductive yarn including a plurality of interlocked fibers; at leastpartially coated with a composition of carbon nanoparticle and SP1 basedpolypeptide (SP1/CNP); one or more polyamine coatings; and an outermetal coating.

According to a further feature of the present invention, wherein thepolyamine coatings are implemented as a first polyamine coatingsandwiched between the fiber and the composition of the SP1/CNP and asecond polyamine coating between the composition and the outer metalcoating.

According to a further feature of the present invention, there is alsoprovided an inner metal coating disposed between the second polyaminecoating and the outer metal coating.

According to a further feature of the present invention, the SP1 basedpolypeptide is non-covalently bound to the carbon nanoparticle.

According to a further feature of the present invention, the SP1 basedpolypeptide is characterized by at least 85% amino acid homology to SEQID NO:1 (wild type); and stable dimer-forming capability.

According to a further feature of the present invention, the SP1 basedpolypeptide is further characterized by at least one conserved aminoacid sequence in at least one region corresponding to amino acids 9-11,44-46 and/or 65-73, of SEQ ID NO:4.

According to a further feature of the present invention, the SP1 basedpolypeptide is a chimeric SP1 polypeptide characterized by at least 85%amino acid homology to SEQ ID NO:1; stable dimer-forming capability; atleast one conserved amino acid sequence in at least one regioncorresponding to amino acids 9-11, 44-46 and/or 65-73, of SEQ ID NO:4;and a carbon surface binding peptide at the N-terminus of the SP1polypeptide.

According to a further feature of the present invention, the carbonsurface binding peptide has an amino acid sequence selected from thegroup consisting of SEQ ID NOs: 10-13.

According to a further feature of the present invention, the SP1 basepolypeptide has the amino acid sequence as set forth in any one of SEQID NOs: 3, 4, 6, 8, 9, 14-18 and 86.

According to a further feature of the present invention, the chimericSP1 polypeptide has the amino acid sequence as set forth in any one ofSEQ ID NOs: 6, 8, 9, 14-18 and 86.

According to a further feature of the present invention, the chimericSP1 polypeptide has the amino acid sequence as set forth in SEQ ID NO:8.

According to a further feature of the present invention, the carbonnanoparticle is implemented as either conductive carbon black,non-conductive carbon black, or carbon nanotube.

According to a further feature of the present invention, the outer metalcoating is implemented as a copper coating.

According to a further feature of the present invention, the inner metalcoating is as Pd(I), Pt(II), Rh(I), Ir(I), iron, aluminum, gold, silver,nickel, or combination thereof.

According to a further feature of the present invention, the fiber isselected from the group consisting of cotton fiber, wool fiber, silkfiber, glass fiber, nylon fiber, polyester fiber, aramid fiber,polyethylene fiber, poly-olefin fiber, polypropylene fiber, and elastanefiber.

According to a further feature of the present invention, wherein theload of the SP1/CNP on the yarn is between 0.01 gr/kg and 100 gr/kg.

According to a further feature of the present invention, the load of theSP1/CNP on the plurality of fibers is between 5 gr/kg and 15 gr/kg.

According to a further feature of the present invention, the load of theSP1/CNP on the plurality of fibers is between 7 gr/kg and 14 gr/kg.

According to a further feature of the present invention, the CNP:SP1ratio is between 0.1:1 to 30:1 dry w/w.

According to a further feature of the present invention, the CNP:SP1ratio is between 2.5:1 to 8:1 dry w/w.

According to a further feature of the present invention, the CNP:SP1ratio is between 5:1 to 7:1 dry w/w.

According to a further feature of the present invention, the yarn has atwist range of at least 10 winds/meter.

According to a further feature of the present invention, the outer metalcoating has a thickness of between 0.01 μm and 100.0 μm.

According to a further feature of the present invention, the yarn has aresistance between 0.001 Ω/m to 1000 mega Ω/m.

According to a further feature of the present invention, the polyamineincludes polyethyleneimine (PEI).

According to a further feature of the present invention, there isfurther provided a polymeric coating on the outer metal coating.

There is also provided according to the teachings of the presentinvention, a method of producing the conductive yarn, includingcontacting a plurality of fibers with a polyamine so as to form aplurality of polyamine coated fibers; contacting the polyamine coatedfibers with a dispersion comprising an SP1/CNP complex so as to form aplurality of complex coated fibers; and contacting the plurality ofcomplex coated fibers with a metal, so as to form plurality of metalcoated fibers.

According to a further feature of the present invention, there is alsoprovided at least one step of washing the yarn with a buffer or waterafter each step of the method.

According to a further feature of the present invention, the each stepof contacting is implemented in a textile dying machine or a bath.

According to a further feature of the present invention, the dispersionincludes an SP1/CNP composition in a concentration of between 0.001% and50% w/w.

According to a further feature of the present invention, the dispersioncomprises an SP1/CNP composition is in a concentration of 0.05%, 0.1%,or between 0.05% and 0.15%.

According to a further feature of the present invention, the dispersioncomprises an SP1/CNP composition is in a concentration of between 1% and20% w/w.

According to a further feature of the present invention, the dispersioncomprises an SP1/CNP composition is in a concentration of 2%, 4%, 6%, or8% w/w.

According to a further feature of the present invention, the PEI isapplied to the yarn at a load of 0.0035% to 30% w/w.

According to a further feature of the present invention, the PEI isapplied to the yarn at a load of 0.007% to 0.7% w/w.

According to a further feature of the present invention, the PEI isapplied to the yarn at a load of about 0.35% w/w.

According to a further feature of the present invention, the method isperformed at a temperature between 4°-90° C.

According to a further feature of the present invention, the metalliccatalyst is implemented as Pd(II), Pt(II), Rh(I), Ir(I), Cu(II) or anycombination thereof.

According to a further feature of the present invention, the metalliccatalyst is implemented as a combination of Pd(II) and Cu(II).

According to a further feature of the present invention, the ratiobetween Pd(II) and Cu(II) is 1:10.

According to a further feature of the present invention, the metalliccatalyst is iron, gold, silver, nickel platinum, aluminum, or anycombination thereof.

According to a further feature of the present invention, there is alsoprovided comprising applying an outer metallic coating of copper outsideof the metallic catalyst coating the fibers.

According to a further feature of the present invention, contacting theplurality of fibers with a metallic catalyst is implemented in a timespan ranging from 5 seconds to 1 hour.

According to a further feature of the present invention, there isfurther provided a step of drying the fibers after contacting the metalcatalyst.

According to a further feature of the present invention, there isfurther provided reducing the metallic catalyst to its metallic state.

According to a further feature of the present invention, the CNP of theSP1/CNP complex is implemented as conductive carbon black (CB_(max)).

According to a further feature of the present invention, there isfurther provided applying a polymeric coating onto the outer metalliccoating of copper.

There is also provided according to the teachings of the presentinvention, conductive yarn including a plurality of interlocked fibersat least partially coated with a composition of carbon nanoparticle andSP1 based polypeptide (SP1/CBmax); one or more polyamine coatings; andan outer metal coating.

According to a further feature of the present invention, there isfurther provided a polymeric coating on the outer metal coating.

According to a further feature of the present invention, the polyaminecoatings are implemented as a first polyamine coating sandwiched betweenthe fiber and the composition of the SP1/CNP and a second polyaminecoating between the composition and the outer metal coating.

There is also provided according to the teachings of the presentinvention, a polymeric film coated at least partially coated with acomposition of carbon nanoparticles (CNP) and SP1 based polypeptide(SP1/CNP); a plurality of polyamine coatings; and a first metal coatingdisposed on the polyamines coating.

According to a further feature of the present invention, the polyaminecoatings are implemented as a first polyamine coating sandwiched betweenthe film and the composition and a second polyamine coating between thecomposition and the first metal coating.

According to a further feature of the present invention, there is alsoprovided a second metal coating disposed between the second polyaminecoating and the first metal coating.

According to a further feature of the present invention, the CNPincludes conductive CB_(max).

BRIEF DESCRIPTION OF THE DRAWINGS

The subject matter regarded as the invention is particularly pointed outand distinctly claimed in the concluding portion of the specification.The invention, however, the invention and its method of production,features, and advantages is best understood in reference to thefollowing detailed description and accompanying drawings in which:

FIGS. 1A and 1B are high resolution scanning electron microscope imagesof SP1/CB (N326) dispersion and polymeric fiber coated with the SP1/CB(N326) complex, respectively;

FIG. 2A is a plot depicting the effect of time of sonication on SP1/CBdispersion at different SP1/CB concentration and dry w/w ratios;

FIG. 2B is a plot depicting suspension as a function of sonication timefor various loading of CB in the SP1/CB complexes;

FIG. 3A is a flow chart depicting steps associated with the productionof conductive yarn;

FIGS. 3B and 3C are schematic, side cross-sectional-views of a dyeingmachine in an in-out and an out-in mode, respectively;

FIG. 4A is a chart depicting electrical resistance as a function of yarnlength under tension;

FIGS. 4B-4D are enlarged images of wound yarn depicting progressivestages of copper covering a carbon black underlayer;

FIGS. 5A-5C are charts depicting electrical resistance as a function ofyard length for low temperature Cu bath at various contact times;

FIGS. 6A-6B are charts depicting electrical resistance as a function ofweight applied to CuSO₄ treated yarn for 10 and 40 minutes,respectively;

FIG. 7A is an image of yarn bobbins after various treatments;

FIGS. 7B-7E are enlarged images of the yarn units of the fully Cutreated bobbin of variety of treatments;

FIG. 8 is a plot depicting electrical resistance as a function oftreatment time in a Cu bath for KEVLAR™ yarn;

FIG. 9A is a plot depicting electrical resistance as a function oftreatment time in a Cu bath for polyester fabric;

FIG. 9B is an image of conductive polyester fabric treated withdifferent palladium concentrations;

FIG. 10A is a plot depicting electrical resistance per length unit as afunction of treatment time in a Cu bath for glass fiber fabric; and

FIGS. 10B-10C are images of glass fiber fabric after various stages oftreatment of in the absence of PEI treatment and in presence of PEItreatment, respectively.

It will be appreciated that for the sake of clarity, elements shown inthe figures have not necessarily been drawn to scale. Further, whereconsidered appropriate, reference numerals may be repeated among thefigures to indicate corresponding or analogous elements.

DETAILED DESCRIPTION OF THE PRESENT INVENTION

In the following detailed description, numerous specific details are setforth in order to provide a thorough understanding of the invention.However, it will be understood by those skilled in the art that thepresent invention may be practiced without these specific details. Inother instances, well-known methods, procedures, and components have notbeen described in detail so as not to obscure the present invention. Itshould be understood that the invention includes combinations offeatures set forth in various embodiments. Also, it is to be understoodthat the phraseology and terminology employed herein is for the purposeof description and should not be regarded as limiting.

The present invention relates to conductive non-metallic polymeric orglass materials having metallic coating and structurally reinforcingSP1/CNP complex providing a platform for bonding of electricallyconductive metals.

The SP1/CNP complex may be implemented as any of several SP1 variantscomplexed with carbon black or carbon nanotubes, or both.

SP1 polypeptide is an exceptionally stable polypeptide, forming hetero-and homo-oligomers which are resistant to denaturation by heat and mostchemical denaturants, resistant to protease digestion, and capable ofstabilizing molecular interactions and forming three dimensionalstructures (Dgany et al, JBC, 2004; 279:51516-23, and U.S. Pat. No.7,253,341 to Wang et al)

SP1 Based Polypeptides

The bi- and multi-functional binding properties of chimeric SP1polypeptides bind to fibers, films and fabrics and can be used to modifythe physical properties of fibers, films and fabrics such as aramid(Kevlar™ and Twaron™), silk, polyester, glass-fiber, polyamide, cotton,nylon, and carbon fibers.

SP1 based polypeptides can be used as a protein scaffold for thepresentation of surface active moieties. A versatile protein scaffoldshould generally constitute a conformationally stable folding entitythat is able to display a multitude of loop structures or amino-acidsequences in a localized surface region. SP1 can be engineered todisplay various moieties contributing to their binding capability in acooperative manner. Moreover, peptide exposure can be manipulated undersolvent conditions that reduce non-specific binding.

Thus, according to one aspect of the present invention the SP1 basedpolypeptide comprised in the composition of matter according to thisinvention is characterized by: i) at least 65% amino acid homology tonative SP1 (SEQ ID NO: 4); and ii) stable dimer-forming capability.

According to another aspect of the present invention the composition ofmatter according to this invention comprises isolated chimericpolypeptides comprising an SP1 polypeptide and a target binding peptide,wherein the SP1 polypeptide is characterized by: i) at least 65% aminoacid homology to native SP1 (SEQ ID NO: 4); ii) stable dimer-formingcapability; and iii) at least one conserved amino acid sequence in atleast one region corresponding to amino acids 9-11, 44-46 and/or 65-73,of SEQ ID NO:4.

As used herein the phrase “SP1 based polypeptide” refers to a proteinhaving at least the following characteristic properties: at least 65%sequence homology to SEQ ID NO:4; and being capable of forming stabledimers. In another embodiment, the phrase refers to a protein having atleast the following characteristic properties: at least 65% sequencehomology to SEQ ID NO:4; being capable of forming stable dimers, andhaving at least one conserved amino acid sequence in at least one regioncorresponding to amino acids 9-11, 44-46 and/or 65-73, of SEQ ID NO:1,as determined using a Best Fit algorithm of GCG, Wisconsin PackageVersion 9.1, using a plurality of 10.00, a threshold of 4, averageweight of 1.00, average match of 2.91 and average mismatch of minus2.00. In some embodiments, the SP1 polypeptide has conserved consensussequences: “HAFESTFES” (65-73, SEQ ID NO:1), “VKH” (9-11, SEQ ID NO:1)and “KSF” (44-46, SEQ ID NO:1). According to one embodiment of theinvention, “wild-type” or “native” SP1 is the stress related SP1 proteinfrom aspen (SEQ ID NO:4), as disclosed by Wang et al (U.S. patentapplication Ser. No. 10/233,409, filed Sep. 4, 2002, now U.S. Pat. No.7,253,341, issued Aug. 7, 2007, which is a Continuation in Part of PCTIL 02/00174, filed Mar. 5, 2002, both of which are incorporated byreference as if fully set forth herein.). The term “SP1 basedpolypeptide” refers in the context of this invention also to chimericSP1 based polypeptide, i.e., polypeptides comprising an SP1 basedpolypeptide and a non-SP1 oligo- or polypeptide. In one embodiment, theSP1 based polypeptide is the crude SP1, i.e., is the heat stablefraction of the crude extract obtained from the bacteria that expressedit. In another embodiment, the SP1 based polypeptide is the pure SP1,obtained by further purification of the heat stable fraction of thecrude extract expressed by the bacteria.

In one embodiment, the SP1 based polypeptide comprised in thecomposition of matter according to this invention is 70%, 75%, 80%, 85%,90%, 95%, or up to 100% homologous to SEQ ID NO: 4. It will beappreciated that SP1 homologues have been identified in plant speciesother than aspen, and that these SP1 homologues can be suitable for usewith the present invention, when fulfilling the abovementioned criteria.

The SP1 based polypeptide comprised in the composition of matteraccording to this invention, can be native SP1 (for example, SEQ ID NO:4), or can be a chimera, i.e., an SP1 polypeptide having a modifiedamino acid sequence. In some embodiments modified SP1 polypeptidesretain the above-mentioned activities of native SP1 polypeptide such asability of forming oligomer and complexes that are pH-stable,hear-stable and denaturant- and protease-resistant.

As mentioned hereinabove, SP1 variant polypeptides can be modified toimpart specific properties to the SP1 variant, thereby rendering themolecular complexing with, and release of other substances moreefficient and controllable, and adaptable to specific conditions. Dganyet al (JBC 2004 279:51516-523) have identified a number of structurallysignificant regions in the SP1 polypeptide.

In some embodiments, the chimeric SP1 polypeptides contemplated in thecomposition of matter of this invention include, but are not limited to,modifications to side chains, incorporation of unnatural amino acidsand/or their derivatives during peptide synthesis and the use ofcross-linkers and other methods which impose conformational constraintson the peptides or their analogues.

According to one embodiment of the present invention any type of SP1based polypeptide, may bind carbon fibers or surfaces. Thus, SP1 wildtype or chimeric polypeptides can be used to bind carbon nanoparticles(e.g., carbon nanotubes, carbon black) and/or graphitic surfaces. Thus,according to further aspects of the present invention there is provideda composition comprising wild type SP1 protein and a carbon nanotube,carbon black, or graphitic surfaces. According to further aspects of thepresent invention there is provided a chimeric polypeptide comprising anSP1 based polypeptide and a carbon nanotube, carbon black, or graphiticsurfaces binding peptide, and a carbon nanotube, carbon black, orgraphitic surface.

According to one embodiment of the present invention the target bindingpeptide binds carbon nanoparticles or carbon surfaces. Thus, thechimeric polypeptides can be used to bind carbon nanoparticles (e.g.,carbon nanotubes, carbon black) and/or graphitic surfaces. Thus,according to further aspects of the present invention there is provideda chimeric polypeptide comprising an SP1 based polypeptide and a carbonnanotube, carbon black, or graphitic surfaces binding peptide. Carbonnanotube binding peptides suitable for use with the chimeric polypeptideof the invention are well known in the art, for example, the peptidesdisclosed in U.S. Pat. No. 7,304,128 to Jagoda et al. According to someembodiments, the carbon nanotube, carbon black or graphitic surfacesbinding peptide is HWSAWWIRSNQS (SEQ ID NO: 10), HSSYWYAFNNKT (SEQ IDNO: 11), DYFSSPYYEQLF (SEQ ID NO:12) or SNQS (SEQ ID NO:13) and thechimeric SP1 based polypeptide has an amino acid sequence as set forthin SEQ ID NOs:6, 8, 9 and 14-18. In certain embodiments, the carbonnanotube, carbon black or graphitic surfaces binding peptide is locatedat the N-terminus of the SP1 polypeptide.

In some embodiments, peptides that non-specifically bind to materialscan be used in the chimeric SP1 based polypeptides comprised in thecomposition of matter according to this the invention. These include,but are not limited to repeated tyrosine rich motifs from specificmussel proteins (mfp1), where the tyrosine residues may be converted toL-DOPA (L-3,4-dihydroxyphenylalanine) (Holten-Andersen & Waite J DentRes 87(8):701-709, 2008), such as AKPSYPPTYK, (SEQ ID NO: 20), AKPTYK(SEQ ID NO: 21), PKISYPPTYK (SEQ ID NO: 22), APPPAXTAXK (SEQ ID NO: 23),ATPKPXTAXK (SEQ ID NO: 24), PYVK (SEQ ID NO: 25), AKPSPYVPTGYK (SEQ IDNO: 26), GQQKQTAYDPGYK (SEQ ID NO: 27).

As used herein, a “chimeric polypeptide” refers to an amino acidsequence having two or more parts which generally are not found togetherin a single amino acid sequence in nature. Chimeric SP1 basedpolypeptides are defined herein as polypeptides comprising an SP1 basedpolypeptide and a non-SP1 oligo- or polypeptide having binding affinityfor target molecules such as carbon nanoparticles (e.g., carbon black,carbon nanotubes, etc.) metals and other ions, the SP1 polypeptide andthe non-SP1 component connected through a peptide bond.

Surprisingly, it was uncovered that SP1-CBD fusion protein binds fibers,fabrics and fabric substrates, as well as to carbon nanotubes with highaffinity. Thus, according to one aspect of the present invention thereis provided a composition of matter comprising an SP1-CBD chimericpolypeptide complexed with carbon nanotubes. According to another aspectthere is provided a composition of matter comprising an SP1-CBD chimericpolypeptide complexed with carbon black.

The SP1-CBD chimeric polypeptide complexed with carbon nanotubes orcarbon black can be used to incorporate carbon nanotubes or carbon blackinto textiles, yarns, fabrics and the like. Thus, in one embodiment,there is further provided an SP1-CBD chimeric polypeptide-CNT-complexedpolymer, fiber, film, fabric or polymeric fabric. In another embodiment,there is further provided an SP1-CBD chimeric polypeptide-CB-complexedpolymer, fiber, film, fabric or polymeric fabric.

The chimeric SP1 polypeptides of the present invention can also be usedto bind carbon nanotube, carbon black and/or graphite surfaces.

Preferred SP1 based polypeptides to be used in composition of matteraccording to this invention include SP1 protein extracts represented by:SEQ ID NO: 4 (WT), SEQ ID NO: 6 (L1-SP1), SEQ ID NO: 14 (L2-SP1), SEQ IDNO: 8 (L3-SP1), SEQ ID NO: 15 (L6-SP1), SEQ ID NO: 3 (mtbSP-SP1), SEQ IDNO: 9 (L4-SP1), SEQ ID NO: 86 (SP1-CBD) or any combination thereof. Inone embodiment, the protein extracts are pure, i.e., undergo a furtherstep of purification after obtaining the heat stable fraction of thecrude extract expressed by the bacteria. In another embodiment, theprotein extracts are crude extracts, i.e., the heat stable fraction ofthe crude extract, used as obtained from the bacteria that expressedthem.

In some embodiments, the SP1 based polypeptide interact with the targetsubstance, or with the chemical environment via a reversible interactionsuch as Van Der Waals (VDW), hydrogen bonds, or electrostaticinteractions, or via non-reversible covalent bonds, all are referred toherein as molecular associations. As used herein the phrase “molecularassociation” refers to a chemical association or a physical associationor both, which takes place on a molecular level. For example, a bond orassociation can be a covalent bond, a VDW interaction, hydrogen bonds,electrostatic interactions, hydrophobic interactions, etc.

Types of reversible molecular associations or bonds suitable for use inthe present invention are associations selected from the groupconsisting of electrostatic bonding, hydrogen bonding, van der Waalsforces, ionic interaction or donor/acceptor bonding. The reversibleassociation can be mediated by one or more associations between thesubstance (i.e., CNP, e.g., CB or CNT) and the SP1 based polypeptide.For example, the reversible association can include a combination ofhydrogen bonding and ionic bonding between the complexing substance andthe SP1 polypeptide. Additionally, or alternatively, the reversibleassociation can be in combination with, for example, covalent or othernoncovalent interactions between components, such as between a substanceand an SP1 based polypeptide or chimeric polypeptide. In one embodiment,the SP1 based polypeptide or the chimeric SP1 based polypeptide interactwith the CNP via hydrophobic interactions. In another embodiment, viahydrogen bonds. In another embodiment, via VDW interactions; or inanother embodiment, via electrostatic interactions. In anotherembodiment, the SP1 based polypeptide or the chimeric SP1 basedpolypeptide interact with the CNP via combination of hydrophobicinteractions, hydrogen bonds, VDW interactions, and electrostaticinteractions. Such interactions between the SP1 based polypeptide or thechimeric SP1 polypeptide and the CNP are also regarded herein as“complexation”, whereas the bound SP1-CNP material is regarded herein asan SP1/CNP “complex”.

The SP1 based polypeptides, chimeric SP1 polypeptides and compositionsof matter comprising the same have been shown to enhance dispersal ofbound substances in a solvent. For example, highly insoluble carbonnanotubes were found to disperse with up to 1000-fold greaterconcentration in both aqueous and organic environments when complexedwith a chimeric, carbon nanotube-binding L1 SP1 (see U.S. Pat. No.8,957,189 herein incorporated by reference in its entirely).

As used herein, the term “dispersion” refers to the ability of a soluteor a colloid to be evenly distributed and/or dissolved in a solvent, inorder to form a solution or suspension comprising the solvent andsolute. It will be appreciated that all solutes are, in theory, solublein all solvents. However, poorly or negligibly soluble (immiscible)solutes or colloids do not form solutions or suspensions of anysignificant concentration with given solvents.

Thus, as used herein, “enhancing the dispersion” refers to increasingthe concentration of said substance, as a solute or colloid, in asolution or suspension with a solvent. In a preferred embodiment, thesubstance is a hydrophobic substance, typically insoluble or poorlysoluble in water, and the solvent is an aqueous solvent.

Stability under centrifugation under standard conditions is used as ameasure for dispersion ability.

The stability of SP1 based polypeptides and chimeric SP1 basedpolypeptides bound to CNT (SP1/CNT complexes) to boiling, proteasedigestion and pH extremes is shown in U.S. Pat. No. 8,957,189.

Following is a list of the ID sequences associated with SP1.

SEQ ID NO. 1 CTGCTCGATCTCATTCCAAGCTGTA AGAGTTTCAATTGGGGCACG SEQ ID NO. 2GCAAGTCTGGTTTGCAAGA GTACTGCGATTCTGCTGCTCTTGCTG SEQ ID NO. 3MRKLPDAATRTPKLVKHTLLTRFKDEITREQIDNYINDYTNLLDLIPSMKSFNWGTDLGMESAELNRGYTHAFESTFESKSGLQEYLDSAALAAFAEGFLPTLSQRLV IDYFLYSEQ ID NO. 4 MATRTPKLVKHTLLTRFKDEITREQIDNYINDYTNLLDLIPSMKSFNWGTDLGMESAELNRGYTHAFESTFESKSGLQEYLDSAALAAFAEGFLPTLSQRLVIDYFLY SEQ ID NO. 5RKLPDAA SEQ ID NO. 6MHWSAWWIRSNQSATRTPKLVKHTLLTRFKDEITREQIDNYINDYTNLLDLIPSMKSFNWGTDLGMESAELNRGYTHAFESIFESKSGLQEYLDSAALAAFAEGFLPT LSQRLVIDYFLYSEQ ID NO. 7 CCACAGAGAG AAAGGGAAGA CATGAAGCTT GTGAAGCACA CATTGTTGACTCGGTTCAAG GATGAGATCA CACGAGAACA GATCGACAAC TACATTAATGACTATACCAA TCTGCTCGAT CTCATTCCAA GCATGAAGAG TTTCAATTGGGGCACGGATC TGGGCATGGA GTCTGCGGAG CTAAACCGAG GATACACTCATGCCTTTGAA TCTACATTTG AGAGCAAGTC TGGTTTGCAA GAGTACCTCGATTCTGCTGC TCTTGCTGCA TTTGCAGAAG GGTTTTTGCC TACTTTGTCACAGCGTCTTG TGATAGACTA CTTTCTCTAC TAA SEQ ID NO. 8MDYFSSPYYEQLFATRTPKLVKHTLLTRFKDEITREQIDNYINDYTNLLDLIPSMKSFNWGTDLGMESAELNRGYTHAFESTFESKSGLQEYLDSAALAAFAEGFLPT LSQRLVIDYFLSEQ ID NO. 9 MHWSAWWIRSNQSATRTPKLVKHTLLTRFKDEITKEQIDNYINDYTNLLDLIPSMKSFNWGTDLGMESAELNRGYTHAFESTFESKSGLQEYLDSAALAAFAEGFLPT LSQRLVIDYFLYSEQ ID NO. 10 HWSAWWIRSNQS SEQ ID NO. 11 HSSYWYAFNNKT SEQ ID NO. 12DYFSSPYYEQLF SEQ ID NO. 13 SNQS SEQ ID NO. 14MHSSYWYAFNNKTATRTPKLVKHTLLTRFKDEITREQIDNYINDYTNLLDLIPSMKSFNWGTDLGMESAELNRGYTHAFESTFESKSGLQEYLDSAALAAFAEGFLPT LSQRLVIDYFLYSEQ ID NO. 15 MSNQSATRTPKLVKHTLLTRFKDEITREQIDNYINDYTNLLDLIPSMKSFNWGTDLGMESAELNRGYTHAFESIFESKSGLQEYLDSAALAAFAEGFLPTLSQRLVID YFLYSEQ ID NO. 16 MHWSAWWIRSNQSATRTPKLVKHTLLTRFKDEITREQIDNYINDYTNLLDLIPSMKSFNWGTDLGMESAELNRGYTHAFESIFESKSGLQEYLDSAALAAFAEGFLPT LSQRLVIDYFLYSEQ ID NO. 17 MHWSAWWIRSNQSATRTPKLVKHTLLTRFKDEICREQIDNYINDYTNLLDLIPSMKSFNWGTDLGMESAELNRGYTHAFESIFESKSGLQEYLDSAALAAFAEGFLPT LSQRLVIDYFLYSEQ ID NO. 18 MHWSAWWIRSNQSATRTPKLVKHTLLTRFKDEITKEQIDNYINDYTNLLDLIPSMKSFNWGTDLGMESAELNRGYTHAFESTFESKSGLQEYLDSAALAAFAEGFLPT LSQRLVIDYFLYSEQ ID NO. 19 RALPDA SEQ ID NO. 20 AKPSYPPTYK SEQ ID NO. 21 AKPTYKSEQ ID NO. 22 PKISYPPTYK SEQ ID NO. 23 APPPAXTAXK SEQ ID NO. 24ATPKPXTAXK SEQ ID NO. 25 PYVK SEQ ID NO. 26 AKPSPYVPTGYK SEQ ID NO. 27GQQKQTAYDPGYK SEQ ID NO. 28ATCCACAGAG AGAAAGGGAA GACATGGCAA CCAGAACTCC AAAGCTTGTGAAGCACACAT TGTTGACTCG GTTCAAGGAT GAGATCACAC GAGAACAGATCGACAACTAC ATTAATGACT ATACCAATCT GCTCGATCTC ATTCCAAGCATGAAGAGTTT CAATTGGGGC ACGGATCTGG GCATGGAGTC TGCGGAGCTAAACCGAGGAT ACACTCATGC CTTTGAATCT ACATTTGAGA GCAAGTCTGGTTTGCAAGAG TACCTCGATT CTGCTGCTCT TGCTGCATTT GCAGAAGGGTTTTTGCCTAC TTTGTCACAG CGTCTTGTGA TAGACTACTT TCTCTACTAAACGCTCAGGA GTAACGACTT CGGCCGGGCT ATTTCATGGT AATAAAGTAATGTAATGTTC AATAAATGCT GGTTTTGAAC CACTGAATGT TCGTGTCTTGATTTCTTGTC TGTGCTAAGT GAAGGGAGTG CTGCTATTCC TTTAAAAATAAAGCCCTTGG GGTTGAGTTG TAGTTTTTCA ATCTTTTTCC CCGATTTATTTCGGTCTTGG TGTTGTT SEQ ID NO. 29*VVKHLVIVQFKEDVTPERLDGLIRGYAGLVDKVPSMKAFHWGTDVSIE Xaa Xaa NMHQGFTHVFESTFESTEGVKEYVYHEFATDFLGSTEKVLIIDF SEQ ID NO. 30*VVKHLVIVQFKEDVTPERLDGLIRGYAGLVDKVPSMKAFHWGTDVSIEN XaaXaa MHQGFTHVFESTFESTEGVKEYVYHPAHVEFATDFLGSTEKVLIIDF SEQ ID NO. 31*VVKHLVIVQFKEDVTPERLEGLIRGYAGLVDKVPSMKAFHWGTDVSIEN XaaXaa MHQGFTHVFESTFESTEGVKEYVYHPAHVEFATDFLGSTEKVLIIDF SEQ ID NO. 32*VVKHILLASEKEEVTQERLDELIRGYAALVGVVPSMKAFHWGTDVSIEN XaaXaa MHQGFTHVFESTFESTEGIKEYIEHPAHVEFAK SEQ ID NO. 33*VVKHILLARFKEDVAPERLDQLIRGYAGLVDLVPSMKAFHWGTDVSIEN XaaXaa MHQGFTHVFESTFESTEGVKEYIEHPAHVEFANEFLPVLEKTLIIDY SEQ ID NO. 34*VVKHLVLARFKEEATPEALD Xaa LIRRYAGLVDAVPSfMKAFHWGTDVTV Xaa Xaa LDTHEGFTHVFESTFESAEGVKEYIAHPSHVEFVDEFLALAEKML IVDY SEQ ID NO. 35MEEAKGPVKHVLLASEKDGVSPEKIEELIKGYANLVNLIEPMKAFHWGKDVSIENLHQGYTHIFESTFESKEAVAEYIAHPAHVEFATIFLGSLDKVLVIDYKPTSVS L SEQ ID NO. 36LHQGYTHILESIFESKEAVAEYIAHPAHVEFATIFLGSLDKVLVIDY SEQ ID NO. 37*VVKHVLLAKFKDDVTPERIEELIKDYANLVNLIPPMKSFHWGKDVSAEN Xaa Xaa LHQGFTHVFESIFESPEGVAEYVAHPAHVEYANLELSCLEKVIVIDY SEQ ID NO. 38*VVKHILLAKEKDGIPPEQIDQLIKQYANLVNLVEPMKAFQWGKDVSIEN XaaXaa LHQGFTHVFESTFDSLEGVAEYIAHPVHVEYANTLLPQLEKFLIVDY SEQ ID NO. 39*HVLLPKLKDYFTPERIELMVDYANLVNLMPRMKSFHSGRDVSAEYLHL Xaa Xaa GCTHVYESTFDSPGVAEYVAHAAHVEYANQDLSCLEKVIAIDY SEQ ID NO. 40MATRTPKLVKHTLATREKDEITREQIDNYINDYTNLLDLIPSMKSENWGTDLGMESAELNRGYTHAFESTFESKSGLQEYLDSAALAAFAEGFLPTLSQRLVIDYFLY SEQ ID NO. 41*KHLCLVRFKEGVVVEDI Xaa Xaa Xaa IEELTKLAAE Xaa LDTVKFFGWGKDVLNQEALTQGFTHVFSMSFASAEDLAAYMGHEKHSAFAATFMAVLDKVVVL DF SEQ ID NO. 42*KHLCLVRFKEGVVVEDI Xaa Xaa Xaa IEELTKLAAELDTVKFFGWGKDVLNQEA Xaa LTQGFTHVESMSFASAEDLAACMGHEKHSAFAATFMAVLDKVVVL DF SEQ ID NO. 43*KHLCMAKFKEGVVVEDI Xaa Xaa Xaa IQELTKLAAELDTVKYFGWGKDVLNQEA Xaa LTQGETHVEVMTFASAEDLAACMGHEKHTAFAATFMAALDKVVVM DF SEQ ID NO. 44*VKHLCLVKFKEEVL Xaa Xaa Xaa VDDILQGMTKLVS EMDMVKSFEWGKDV Xaa LNQEMLTQGFTHVFSLTFASSEDLTTYMSHERHQEFAGTFMAAIDKVV VVDFSEQ ID NO. 45* RRPTMGEVKHLCLVKFKEGVVVEDVLKGMTDLVAGMDMV Xaa Xaa Xaa KSFEWGQDV Xaa LNQEMLTQGFTHVFSLTFAFADDLATYMGHDRHAAFAATFMA ALDKVVVIDFSEQ ID NO. 46* ESTFESTEGIKEYIEHPAHVEFAK Xaa LNQEMLTQGFTHVFSLTFATAADLAAYMAHDSHTAFAATFMAAIDKVLVVDF SEQ ID NO. 47*KHLVLVKFKEDVVVEDILKELEKLVQEMDIV Xaa Xaa Xaa KSFVWGKDV Xaa Xaa ESHEMLRQGFTHAIIMTENSKEDYQTFANHPNHVGFSATFATVIDK AVLLDFSEQ ID NO. 48* LLVKFKQDVVEEDVLKQIEQLVNEIDLI Xaa Xaa Xaa KSFVWGKDT Xaa Xaa ESNEMVTQGYTHAMIMTENSKEDYEACVVKEV Xaa Xaa EFSAIFVTV VEKILVLNFSEQ ID NO. 49* HYVIVKFKDGVA Xaa Xaa Xaa VDDLIQGLEKMVEGIDHVKSFEWGKDIXaa Xaa ESHDMLRQGFTHAFLMTFNGKEEFNAFQTHPNHLEFSGVFSPAIEK IVVLDFSEQ ID NO. 50* HYVIVKFKDGVA Xaa Xaa Xaa VDELIQGLEKMVSGIDHVKSFEWGKDI Xaa Xaa ESHDMLRQGFTHVFLMAFNGKEEFNAFQTHPNHLEFTGVFSPAIEK IVVLDFSEQ ID NO. 51* KHEVIVKFKEGVA Xaa Xaa Xaa VDELTKGMEKLVTEIGAVKSFEWGQDI Xaa Xaa ESLDVLRQGFTHAFLMTFNKKEDFVAFQSHPNHVEFSTKFSAAIEN IVLLDFSEQ ID NO. 52* LVSEIHAVKSFEWGQDI Xaa Xaa ESLDVLRQGFTHAFLMTFNKKRRLSEQ ID NO. 53 MATSGEKHLVVVKFKEDTKVDEILKGLENLVSQIDTVKSFEWGEDKESHDMLRQGFTHAFSMIPENKDGYVAFTSHPLHVEFSAAFTAVIDKIVLLDFPVAAVKSSVV ATPSEQ ID NO. 54* KTVEHIVLFKVKEETEPSKVSDMVNGLGSLVSLDPVLH Xaa LSVGPLLRNRSSALT Xaa Xaa FTHMLHSRYKSKEDLEAYSAHPSHVSVVKGYVLPIIDDIMS VDW SEQ ID NO. 55AAAACATATG CGCAAACTTC CGGATGCGGC AACCAGAACT CCAAAGCTTGTGAAGCACAC ATTGTTGACT CGGTTCAAGG ATGAGATCAC ACGAGAACAGATCGACAACT ACATTAATGA CTATACCAAT CTGCTCGATC TCATTCCAAGCATGAAGAGT TTCAATTGGG GCACGGATCT GGGCATGGAG TCTGCGGAGCTAAACCGAGG ATACACTCAT GCCTTTGAAT CTACATTTGA GAGCAAGTCTGGTTTGCAAG AGTACCTCGA TTCTGCTGCT CTTGCTGCAT TTGCAGAAGGGTTTTTGCCT ACTTTGTCAC AGCGTCTTGT GATAGACTAC TTTCTCTACT AA SEQ ID NO. 56AAGGAGATAT ACAAAAACAT ATGCACTGGT CAGCATGGTG GATACGATCAAATCAATCAG CAACCAGAAC TCCAAAGCTT GTGAAGCACA CATTGTTGACTCGGTTCAAG GATGAGATCA CACGAGAACA GATCGACAAC TACATTAATGACTATACCAA TCTGCTCGAT CTCATTCCAA GCATGAAGAG TTTCAATTGGGGCACGGATC TGGGCATGGA GTCTGCGGAG CTAAACCGAG GATACACTCATGCCTTTGAA TCTACATTTG AGAGCAAGTC TGGTTTGCAA GAGTACCTCG ATTCTGCTGC TCTTGCTGCA TTTGCAGAAG GGTTTTTGCC TACTTTGTCA CAGCGTCTTG TGATAGACTA CTTTCTCTAC TAA SEQ ID NO. 57GAAGGAGATA TACAAAAACA TATGCACTCA TCATACTGGT ACGCATTCAACAACAAAACA GCAACCAGAA CTCCAAAGCT TGTGAAGCAC ACATTGTTGACTCGGTTCAA GGATGAGATC ACACGAGAAC AGATCGACAA CTACATTAATGACTATACCA ATCTGCTCGA TCTCATTCCA AGCATGAAGA GTTTCAATTGGGGCACGGAT CTGGGCATGG AGTCTGCGGA GCTAAACCGA GGATACACTCATGCCTTTGA ATCTACATTT GAGAGCAAGT CTGGTTTGCA AGAGTACCTCGATTCTGCTG CTCTTGCTGC ATTTGCAGAA GGGTTTTTGC CTACTTTGTC ACAGCGTCTT GTGATAGACT ACTTTCTCTA CTAA SEQ ID NO. 58ATACAAAAAC ATATGGATTA TTTTTCATCA CCATATTATG AACAATTATT TGCAACCAGA ACTCCAAAGC TTGTGAAGCA CACATTGTTG ACTCGGTTCA AGGATGAGAT CACACGAGAA CAGATCGACA ACTACATTAA TGACTATACC AATCTGCTCG ATCTCATTCC AAGCATGAAG AGTTTCAATT GGGGCACGGA TCTGGGCATG GAGTCTGCGG AGCTAAACCG AGGATACACT CATGCCTTTG AATCTACATT TGAGAGCAAG TCTGGTTTGC AAGAGTACCT CGATTCTGCT  TACTTTCTCT ACTAASEQ ID NO. 59 AGAAGGAGAT ATACAAAAAC ATATGTCAAA TCAATCAGCA ACCAGAACTCCAAAGCTTGT GAAGCACACA TTGTTGACTC GGTTCAAGGA TGAGATCACACGAGAACAGA TCGACAACTA CATTAATGAC TATACCAATC TGCTCGATCTCATTCCAAGC ATGAAGAGTT TCAATTGGGG CACGGATCTG GGCATGGAGTCTGCGGAGCT AAACCGAGGA TACACTCATG CCTTTGAATC TACATTTGAG AGCAAGTCTG GTTTGCAAGA GTACCTCGAT TCTGCTGCTC TTGCTGCATTTGCAGAAGGG TTTTTGCCTA CTTTGTCACA GCGTCTTGTG ATAGACTACT  TTCTCTACTA ASEQ ID NO. 60 AAGGAGATAT ACAAAAACAT ATGCACTGGT CAGCATGGTG GATACGATCAAATCAATCAG CAACCAGAAC TCCAAAGCTT GTGAAGCACA CATTGTTGACTCGGTTCAAG GATGAGATCA CAAAAGAACA GATCGACAAC TACATTAATGACTATACCAA TCTGCTCGAT CTCATTCCAA GCATGAAGAG TTTCAATTGGGGCACGGATC TGGGCATGGA GTCTGCGGAG CTAAACCGAG GATACACTCATGCCTTTGAA TCTACATTTG AGAGCAAGTC TGGTTTGCAA GAGTACCTCGATTCTGCTGC TCTTGCTGCA TTTGCAGAAG GGTTTTTGCC TACTTTGTCA CAGCGTCTTG TGATAGACTA CTTTCTCTAC TAA SEQ ID NO. 61AAGGAGATAT ACAAAAACAT ATGCACTGGT CAGCATGGTG GATACGATCAAATCAATCAG CAACCAGAAC TCCAAAGCTT GTGAAGCACA CATTGTTGACTCGGTTCAAG GATGAGATCT GCCGAGAACA GATCGACAAC TACATTAATGACTATACCAA TCTGCTCGAT CTCATTCCAA GCATGAAGAG TTTCAATTGGGGCACGGATC TGGGCATGGA GTCTGCGGAG CTAAACCGAG GATACACTCATGCCTTTGAA TCTACATTTG AGAGCAAGTC TGGTTTGCAA GAGTACCTCGATTCTGCTGC TCTTGCTGCA TTTGCAGAAG GGTTTTTGCC TACTTTGTCA CAGCGTCTTG TGATAGACTA CTTTCTCTAC TAA SEQ ID NO. 62AAGGAGATAT ACAAAAACAT ATGCACTGGT CAGCATGGTG GATTCGTTCAAATCAATCAG CAACCAGAAC TCCAAAGCTT GTGAAGCACA CATTGTTGACTCGGTTCAAG GATGAGATCA CACGAGAACA GATCGACAAC TACATTAATGACTATACCAA TCTGCTCGAT CTCATTCCAA GCATGAAGAG TTTCAATTGGGGCACGGATC TGGGCATGGA GTCTGCGGAG CTAAACCGAG GATACACTCATGCCTTTGAA TCTACATTTG AGAGCAAGTC TGGTTTGCAA GAGTACCTCGATTCTGCTGC TCTTGCTGCA TTTGCAGAAG GGTTTTTGCC TACTTTGTCA CAGCGTCTTG TGATAGACTA CTTTCTCTAC TAA SEQ ID NO. 63AAGGAGATAT ACAAAAACAT ATGCACTGGT CAGCATGGTG GATTCGTTCAAATCAATCAG CAACCAGAAC TCCAAAGCTT GTGAAGCACA CATTGTTGACTCGGTTCAAG GATGAGATCA CAAAAGAACA GATCGACAAC TACATTAATGACTATACCAA TCTGCTCGAT CTCATTCCAA GCATGAAGAG TTTCAATTGGGGCACGGATC TGGGCATGGA GTCTGCGGAG CTAAACCGAG GATACACTCATGCCTTTGAA TCTACATTTG AGAGCAAGTC TGGTTTGCAA GAGTACCTCGATTCTGCTGC TCTTGCTGCA TTTGCAGAAG GGTTTTTGCC TACTTTGTCA CAGCGTCTTG TGATAGACTA CTTTCTCTAC TAA SEQ ID NO. 64MKLVKHTLLTREKDEITREQIDNYINDYTNLLDLIPSMKSENWGTDLGMESAELNRGYTHAFESTEESKSGLQEYLDSAALAAFAEGELPTLSQRLVIDYELY SEQ ID NO. 65CTGCTCGATCTCATTCCAAGCTGTA AGAGTTTCAATTGGGGCACG SEQ ID NO. 66GCAAGTCTGGTTTGCAAGA GTACTGCGATTCTGCTGCTCTTGCTG SEQ ID NO. 67AAAACATATGCGCAAACTTCCGGATGCGGCAACCAGAACTCCAAAGCTTG SEQ ID NO. 68AAAAGAGCTCTTAGTAAAGAAAGTAATCAATAAC SEQ ID NO. 69ATGAAGCTTG TGAAGCACAC ATTGTTGACT CGGTTCAAGG ATGAGATCACACGAGAACAG ATCGACAACT ACATTAATGA CTATACCAAT CTGCTCGATCTCATTCCAAG CTGTAAGAGT TTCAATTGGG GCACGGATCT GGGCATGGAGTCTGCGGAGC TAAACCGAGG ATACACTCAT GCCTTTGAAT CTACATTTGA GAGCAAGTCT GGTTTGCAAG AGTACCTCGA TTCTGCTGCT CTTGCTGCAT TTGCAGAAGG GTTTTTGCCT ACTTTGTCAC AGCGTCTTGT GATAGACTAC TTTCTCTACT AASEQ ID NO. 70 AAGGAGATATACAAAAACATATGCACTGGTCAGCATGGTGGATACGATCAAATCAATCAGCAACCAGAACTCCAAAG SEQ ID NO. 71CTTTGGAGTTCTGGTTGCTGATTGATTTGATCGTATCCACCATGCTGACCAGTGCATATGTTTTTGTATATCTCCTT SEQ ID NO. 72AGAAGGAGATATACAAAAACATATGCACTCATCATACTGGTACGCATTCAACAACAAAACAGCAACCAGAACTCCAAAGC SEQ ID NO. 73GCTTTGGAGTTCTGGTTGCTGTTTTGTTGTTGAATGCGTACCAGTATGATGAGTGCATATGTTTTTGTATATCTCCTTCT SEQ ID NO. 74ATACAAAAACATATGGATTATTTTTCATCACCATATTATGAACAATTATTTG CAACCAGAACTCCSEQ ID NO. 75 GGAGTTCTGGTTGCAAATAATTGTTCATAATATGGTGATGAAAAATAATCCATATGTTTTTGTAT SEQ ID NO. 76AGAAGGAGATATACAAAAACATATGTCAAATCAATCAGCAACCAGAACTC  CAAAGC SEQ ID NO. 77GCTTTGGAGTTCTGGTTGCTGATTGATTTGACATATGTTTTTGTATATCTCCTT  CT SEQ ID NO. 78ACTGGTCAGCATGGTGGATTCGATCAAATCAATCAG SEQ ID NO. 79CTGATTGATTTGATCGAATCCACCATGCTGACCAGT SEQ ID NO. 80GTCAGCATGGTGGATTCGTTCAAATCAATCAGCAACC SEQ ID NO. 81GGTTGCTGATTGATTTGAACGAATCCACCATGCTGAC SEQ ID NO. 82TGACTCGGTTCAAGGATGAGATCACAAAAGAACAGATCGACA SEQ ID NO. 83TGTCGATCTGTTCTTTTGTGATCTCATCCTTGAACCGAGTCA SEQ ID NO. 84ACTCGGTTCAAGGATGAGATCTGCCGAGAACAGATCGACAACTAC SEQ ID NO. 85GTAGTTGTCGATCTGTTCTCGGCAGATCTCATCCTTGAACCGAGT SEQ ID NO. 86MAATSSMSVEFYNSNKSAQTNSITPIIKITNTSDSDLNLNDVKVRYYYTSDGTQGQTFWCDHAGALLGNSYVDNTSKVTANFVKETASPTSTYDTYVEFGFASGRATLKKGQFITIQGRITKSDWSNYTQTNDYSFDASSSTPVVNPKVTGYIGGAKVLGTAPAVPSGSVTSTSKTTTTASKTSTSTSSTSEFMATSTPKLVKHTLLTRFKDEITREQIDNYINDYTNLLDLIPSMKSFNWGTDLGMESAELNRGYTHAFESTFESKSGLQEYLDSAALAAFAEGFLPTLSQRLVIDYFLY SEQ ID NO. 87MAATSSMSVEFYNSNKSAQTNSITPIIKITNTSDSDLNLNDVKVRYYYTSDGTQGQTFWCDHAGALLGNSYVDNTSKVTANFVKETASPTSTYDTYVEFGFASGRATLKKGQFITIQGRITKSDWSNYTQTNDYSFDASSSTPVVNPKVTGYIGGAKVLGTA P SEQ ID NO. 88MATSTPKLVKHTLLTRFKDETTREQIDNYINDYTNLLDLIPSMKSFNWGTDLGMESAELNRGYTHAFESTFESKSGLQEYLDSAALAAFAEGFLPTLSQRLVIDYFLY SEQ ID NO. 89AVPSGSVTSTSKTTTTASKTSTSTSSTSEF

In some embodiments, the SP1 polynucleotide sequence is 70%, 75%, 80%,85%, 90%, 95%, or up to 100% homologous to SEQ ID NO: 28. It will beappreciated that polynucleotides encoding SP1 homologues SEQ ID NOs:29-54 can be suitable for producing the SP1 polypeptide of the presentinvention, when fulfilling the abovementioned criteria.

SP1/Carbon Nanoparticle

This invention describes compositions comprising SP1 Based polypeptide,and carbon nanoparticles (CNPs) such as: carbon black (CB) and carbonnanotubes (CNT) (referred herein as “SP1/CNP” composition).

According to this invention, the term “carbon nanoparticle” refers bothto carbon nanotubes and to carbon black.

The SP1/CNP compositions are preferably obtained as dispersion in asolvent. The solvent used for the dispersion can be water, commonorganic solvents or a mixture thereof. Non-limiting exemplary organicsolvents include less polar hydrocarbon solvent, such as pentanes,hexanes, petroleum ether, benzene and toluene; and polar solvents, suchas ether, tetrahydrofuran, dichloromethane, chloroform, dichloroethane,dimethylsulfoxide, dimethylformamide, dimethylacetamide, dioxane,methanol, ethanol ethyl acetate, acetonitrile, acetone and carbontetrachloride. Preferably, the solvent is an aqueous solution. Morepreferably the solvent is water.

As used herein, “films” refer to polymeric sheet. Plastic films have awide variety of applications including packaging, plastic bags, labels,building construction, landscaping, electrical fabrication, printingsubstrates photographic film, film stock for movies, video tape, etc.Common plastic films include but are not limited to: Polyethylene film(Low-density polyethylene, Medium-density polyethylene, High-densitypolyethylene, and Linear low-density polyethylene), Polypropylene film(a cast film, biaxially oriented film (BOPP), or a uniaxially orientedfilm), Polyester film (BoPET is a biaxially oriented polyester film),Nylon film, Polyvinyl chloride film (with or without a plasticizer), anda variety of bioplastics and biodegradable plastic films.

According to some embodiments of the present invention the SP1 basedpolypeptides, and chimeric SP1-carbon surfaces (i.e. CNT or graphiticsurfaces) polypeptides can bind to carbon nanoparticles, formingcomposition which can further bind to polymer fibers such as carbonfibers, polyesther, kevlar, nylon, cotton, wool and the like, and can beused to modify the properties of such fibers. For example, the SP1/CB,SP1/CNT/can bind to polymer fibers to facilitate to provide addedstrength, or can be used to promote binding of carbon nanoparticles tosynthetic or natural fabrics, fabric precursors and films, such aspolyester, aramid (Kevlar™), nylon or cotton, in a uniform manner, toprovide fibers and fabrics that have unique chemical, electrical, andthermal properties. Such fabrics and surfaces may comprise layerscomprising carbon nanoparticle surfaces associated with certainpolymeric substances and resins.

According to some embodiments of the present invention, there areprovided compositions of matter comprising an SP1 based polypeptide ofthe present invention and carbon nanoparticle (e.g., CNT, CB), having atarget binding peptide component, and the target substance. Such acomposition of matter can include, in some embodiments, for example,L1SP1 chimera (SEQ ID NO: 6) bound to carbon nanotube, L1SP1 (SEQ ID NO:6) bound to carbon fibers, L3SP1 (SEQ ID NO: 8) bound to carbon black,L3SP1 (SEQ ID NO: 8) bound to carbon nanotube, and the like. In anotherembodiment, the composition of matter according to this invention maycomprise a non-chimeric SP1 based polypeptide, i.e., without a targetbinding peptide component. e.g., SP1 wild-type, while still comprising atarget substance such as carbon nanoparticle (e.g., CNT, CB).

As used herein, the phrase “SP1 based polypeptide-CNP complex” or“SP1/CNP” refers to a composition of matter comprising an SP1 orchimeric SP1 based polypeptide, bound to at least one carbonnanoparticle. In one embodiment, the carbon nanoparticle is carbonblack. In another embodiment the carbon nanoparticle is carbon nanotube.In another embodiment, the composition is in the form of dispersion. Ina preferred embodiment, the dispersion is an aqueous dispersion.

In one embodiment, this invention is directed to a polymer, fiber, film,fabric or polymeric fabric coated with, or complexed to a composition ofmatter comprising SP1 based polypeptide, CNP as described herein above.In another embodiment, the polymer, fiber, film, fabric or polymericfabric comprises at least one of: cotton, wool, silk, nylon, polyester,polypropylene, glass-fiber, elastane, Kevlar and aramid. In anotherembodiment, the polymer, fiber, film, fabric or polymeric fabric ispolyester. In another embodiment, the polymer, fiber, film, fabric orpolymeric fabric is Aramid. In another embodiment, the polymer, fiber,film, fabric or polymeric fabric is nylon. In another embodiment, thenylon is a nylon fabric. In another embodiment the nylon is a nylonfilm.

As used herein, the phrase “SP1 polypeptide-CNP-polymer complex”, alsoregarded as “SP1/CNP coated fiber” or “SP1/CNP enforced fiber” refers toa polymer, fiber, yarn, cord, film or fabric coated with, or complexedto a composition comprising an SP1 based or chimeric SP1 basedpolypeptide and CNP bound thereto. In one embodiment, the CNP is carbonblack. In another embodiment the CNP is carbon nanotube.

In another embodiment, the polymer, fiber, film, fabric or polymericfabric according to this invention further comprises a polyethyleneimine(PEI). In another embodiment, the polymer, fiber, film, fabric orpolymeric fabric according to this invention is coated with at least onelayer of polyethyleneimine (PEI). In another embodiment, with one layer;two layers; three layers; or four layers of PEI; each possibilityrepresents a separate embodiment of the invention.

“Polyethylenimine” (PEI) (also called “polyaziridine”) is a polymer withrepeating unit composed of the amine group and two carbon aliphaticCH₂CH₂ spacer. Linear polyethyleneimines contain all secondary amines,in contrast to branched PEIs which contain primary, secondary andtertiary amino groups. PEI is available at various molecular weights. Inone embodiment, the PEI that finds utility in the context of thisinvention has a molecular weight of between 800 Da and 750,000 Da. Inanother embodiment, the PEI is high molecular weight PEI. In anotherembodiment the PEI is low molecular weight PEI. In another embodiment,the molecular weight of the PEI is between 1 KDa and 10,000 KDa; Inanother embodiment, between 10 KDA and 100 KDa; In another embodiment,between 25 KDa and 80 KDa; In another embodiment, between 50 KDa and 70KDa. In another embodiment, the molecular weight of the PEI is about 60KDa.

In another embodiment, the loading of the PEI applied to the fiber, filmor fabric is between about 0.05% and about 5% (weight PEI/weightfabric). In another embodiment, between about 0.5% and about 2.5%. Inanother embodiment, between about 0.5% and about 1.5%. In anotherembodiment, between about 1.8% and about 2.0%. In another embodiment,between about 0.1% and about 1.0%.

In one embodiment, the fiber, yarn, cord, film, or fabric coated withthe composition of matter according to this invention comprises a wovenor non-woven fiber, yarn, cord, film, or fabric. In another embodimentsaid woven and non-woven fiber, yarn, cord, film, or fabric is selectedfrom natural fiber, yarn, cord, film, or fabric, synthetic fiber, yarn,cord, film, or fabric, a mixture of natural and synthetic fiber, yarn,cord, film, or fabric, and inorganic material based fiber, yarn, cord,film, or fabric. Exemplary natural fabrics include, but are not limitedto cotton, wool and silk. Exemplary synthetic fabric fiber or filminclude, but are not limited to nylon, polyester, aramid, rayon,polypropylene, polyethylene naphthanate (PEN), polyolefin ketone (POK),and elastane (Lycra™-Spandex™). Examples of garments, rope, sewn, moldedand woven items fashioned from fabric and yarns coated with the SP1/CNTaccording to the present invention include, but are not limited to:parachutes, clothing, sleeping bags, bicycle parts and equipment, skis,etc (for further detailed examples, see U.S. Pat. Nos. 7,354,877, and8,957,189, which are incorporated herein by reference).

In one embodiment, the polymer, fiber, film, fabric or polymeric fabricof this invention comprises a plurality of layers of the composition ofmatter of this invention bound to said polymer, fiber, film, fabric orpolymeric fabric. In another embodiment, it comprises one layer of saidcomposition of matter bound to said polymer, fiber, film, fabric orpolymeric fabric. In another embodiment, it comprises two layers of saidcomposition of matter bound to said polymer, fiber, film, fabric orpolymeric fabric. In another embodiment, it comprises three layers ofsaid composition of matter bound to said polymer, fiber, film, fabric orpolymeric fabric.

In one embodiment, this invention is directed to a polymer, fiber, film,fabric or polymeric fabric coated with at least one layer of acomposition of matter comprising SP1 based polypeptide according to thisinvention, Carbon nanoparticle (i.e., SP1/CB, SP1/CB). In one embodimentthe fiber, film or fabric is coated with at least one layer of SP1/CB.In another embodiment, the fiber, film or fabric is coated with at leastone layer of SP1/CB/metal. In another embodiment, the fiber, film orfabric is coated with at least one layer of SP1/CNT/metal.

In one embodiment, this invention is directed to a polymer, fiber, film,fabric or polymeric fabric complexed with a composition of mattercomprising SP1 based polypeptide according to this invention, Carbonnanoparticle (i.e., SP1/CB). In one embodiment the fiber, film or fabricis complexed with SP1/CB.

In another embodiment the carbon nanoparticle (CNP) is carbon nanotube.

Further embodiments of the invention include, but are not limited to,tires having at least one SP1 based polypeptide-CNT-complexed polymer,fiber, film, fabric or polymeric fabric element forming a conductivepath for discharging static electric charge buildup (for details ofconstruction of such tires, see, for example, US Patent Application2010078103, to Nakamura, U.S. Pat. No. 7,528,186 to Halas a, U.S. Pat.No. 7,284,583 to Dheur et al and U.S. Pat. No. 7,131,474 to Sandstrom),tires having at least one SP1 based polypeptide-CNT-complexed polymer,fabric or polymeric fabric element having improve thermal conductivityand heat transfer to the road during use (for details of constructionand use of such tires, see, for example, U.S. Pat. No. 7,337,815 toSpadone), tires having at least one SP1 based polypeptide-CNT-complexedpolymer, fabric or polymeric fabric element capable of communicatinginformation on tire status and performance, such as tire pressure,deformation, tread and sidewall wear and failure, rolling resistance,etc. at rest and in motion, during use (for details of construction anduse of such tires, see, for example, U.S. Pat. No. 7,318,464 to Hahn etal and U.S. Pat. No. 7,581,439 to Rensel, et al.) and electrical energygenerating tires with a conductive strip of a SP1 basedpolypeptide-CNT-complexed polymer, fabric or polymeric fabric, and anenergy generating component (such as a piezo-ceramic orthermal-harvesting material) incorporated into the tread and/or sidewallof the tire ((for details of construction and use of such tires, see,for example, U.S. patent application Ser. No. 00/70,028,958 to Retti andU.S. patent application Ser. No. 00/90,314,404 to Rodgers et al).

SP1 based polypeptides complexed with target substances can be added topolymers, fibers, films, or fabrics in a variety of methods. In someembodiments, the complex of SP1 based polypeptide and target substance(for example, L3-SP1-CB complex) is prepared similarly to the methoddescribed in U.S. Pat. No. 8,957,189 for SP1-CNT complexes.Alternatively, the complex of SP1 based polypeptide and target substance(e.g. carbon nanotube, carbon black) is prepared by as described hereinbelow.

This invention also relate to methods of producing fibers, yarns, cords,films, fabrics or polymeric fabrics enforced with carbon nanoparticles(CNP). In one embodiment, the method for producing such fibers, yarns,cords, films, fabrics or polymeric fabrics comprise contacting adispersion comprising composition of matter comprising SP1 basedpolypeptide according to this invention and CNP (SP1/CNP), with apolymer, fiber, film, fabric or polymeric fabric. Preferably, the methodcomprises a pre-step of de-sizing of said fibers, yarns, cords, films,fabrics or polymeric fabrics prior to contacting it with thecomposition. In one embodiment, the method further comprises a step ofwashing the unbound composition from said polymer, fiber, film, fabricor polymeric fabric with buffer or water. In another embodiment, themethod further comprises a step of contacting the polymer, fiber, film,fabric or polymeric fabric, with polyethyleneimine (PEI) prior tocontacting the polymer, fiber, film, fabric or polymeric fabric withsaid composition, and after the de-sizing pre-step. Preferably, thesteps of contacting the fiber with PEI, contacting the dispersion withthe fiber, and washing the unbound composition, are repeated at leastonce; in another embodiment, they are repeated twice; three, four times,or even up to ten times. In one embodiment, the method further comprisesa post-treatment step of contacting the fiber, yarn, cord, film, fabricor polymeric fabric with PEI as the last step. In one embodiment, theCNP is implemented as CB. In another embodiment, the CNP is implementedas CNT. In another embodiment, said step of contacting the dispersioncomprising the composition of this invention with the polymer, fiber,film, or fabric, is performed using a textile dying machine. Nonlimiting examples of textile dying machines are: Jigger coating machinefor woven fabrics or vertical or horizontal yarn or fabric packagedyeing system.

In one embodiment, a method of producing a fiber, yarn, cord, film,fabric or polymeric fabric coated with carbon nanoparticles includes:

-   -   a. Optionally de-sizing a fiber, yarn, cord, film, fabric or        polymeric fabric;    -   b. Optionally contacting said fiber, yarn, cord, film, fabric or        polymeric fabric with polyethyleneimine (PEI);    -   c. Contacting a dispersion comprising a composition of SP1 Based        polypeptide and CNP according to this invention with the fiber,        yarn, cord, film, fabric or polymeric fabric; and    -   d. Optionally repeating steps (b) and (c) at least once;    -   e. Optionally contacting said fiber, yarn, cord, film, fabric or        polymeric fabric, with polyethyleneimine (PEI).

In another embodiment, the method further comprises a step of washingthe fiber, yarn, cord, film, fabric or polymeric fabric with a buffer orwater after each step.

In another embodiment, said step of contacting the dispersion comprisingthe composition of this invention with the polymer, fiber, film, orfabric, is performed using a textile dying machine.

SP1/Carbon Black (CB)

The present invention provides, in some embodiments thereof, the abilityto weave carbon black into fibers, films and fabrics that may be appliedto a wide range of uses.

Unexpectedly, it was found by the inventors that SP1 based polypeptidesare capable of enhancing the dispersion of not only CNT; but also, CB inaqueous solutions. Such SP1/CB dispersions can be utilized for thereinforcement of polymer fibers, fabrics and films.

CB binding to fibers via the SP1 protein provides additional surface forreceiving metallic conductors, improves interaction with the fibers, andinduces cross linking between the fibers. In addition protein binding tothe fiber by itself may improve the interaction with the polymer throughreactive groups on the protein surface. It is demonstrated that SP1variants according to this invention, are capable of binding CB tostructural fibers.

Successful incorporation of CB into the fabric allows the fabrics toadopt some of the mechanical, thermal, electrical, physical and chemicalproperties associated with carbon black. The SP1/CB compositions ofmatter according to this invention can be utilized to incorporate the CBinto the fabrics.

“Carbon black” (CB) is a form of para-crystalline carbon that has a highsurface-area-to-volume ratio, albeit lower than that of activatedcarbon. It is dissimilar to soot in its much highersurface-area-to-volume ratio and significantly lower (negligible andnon-bioavailable) PAH (polycyclic aromatic hydrocarbon) content. It isproduced by the incomplete combustion of heavy petroleum products suchas FCC tar, coal tar, ethylene cracking tar, and a small amount fromvegetable oil. It is used often in the Aerospace industry in elastomersfor aircraft vibration control components such as engine mounts. In thecontext of this invention, any type of CB can be used. The various CBtypes and brands are differing in their surface area, aggregate sizedistribution and morphological structure. Non limiting examples of CBtypes include: conductive CB, CB-N326, CB-N220, CB-N660, CRX™ 1346, CRX™1490, PROPEL™ D11, PROPEL™ E3, PROPEL™ E6, PROPEL™ E7, REGAL® 300,VULCAN® 10, VULCAN® 10H, VULCAN® 10HD, VULCAN® 1345, VULCAN® 1380,VULCAN® 1391, VULCAN® 1436, VULCAN® 3, VULCAN® 3H, VULCAN® 6, VULCAN®6-LP, VULCAN® 7H, VULCAN® 8, VULCAN® 9, VULCAN® 9H, VULCAN® J, VULCAN®M. In general CB particles ranging from 10 nm to 10 micron may be used.

Accordingly provided herein is a composition of matter comprising carbonblack (CB) bound to an SP1 based polypeptide according to this invention(referred herein as “SP1/CB” composition). In another embodiment, thecarbon black is non-covalently bound to the SP1 based polypeptide. Inanother embodiment, the SP1 based polypeptide is a wild type SP1protein. In another embodiment, the SP1 based polypeptide is a chimericSP1 based polypeptide as described herein above. In another embodiment,the SP1 based polypeptide comprises a carbon nanotube or graphiticsurfaces binding peptide. In another embodiment, the chimeric SP1polypeptide has the amino acid sequence as set forth in SEQ ID NO: 8.

Accordingly, in another embodiment, provided herein is a composition ofmatter comprising carbon black (CB), SP1 based polypeptide according tothis invention. In one embodiment, the CB is non-covalently bound to theSP1 based polypeptide.

The SP1/CB compositions of matter according to this invention arepreferably obtained as dispersions in solvent. The solvent used for thedispersion can be water, common organic solvents or a mixture thereof.Non-limiting exemplary organic solvents include less polar hydrocarbonsolvent, such as pentanes, hexanes, petroleum ether, benzene andtoluene; and polar solvents, such as ether, tetrahydrofuran,dichloromethane, chloroform, dichloroethane, dimethylsulfoxide,dimethylformamide, dimethylacetamide, dioxane, methanol, ethanol, ethylacetate, acetonitrile, acetone and carbon tetrachloride. Preferably, thesolvent is an aqueous solution. More preferably the solvent is water.

In one embodiment, the SP1/CB is in the form of aqueous dispersions in aconcentration of between 0.01% and 50% w/w. In another embodiment, thecomposition is in the form of an aqueous dispersion in a concentrationof between 0.5% and 30%. In another embodiment, the composition is inthe form of an aqueous dispersion in a concentration of between 1% and20%. In another embodiment, the composition is in the form of an aqueousdispersion in a concentration of between 4% and 17%. In anotherembodiment, the composition is in the form of an aqueous dispersion in aconcentration of 1% w/w, 2% w/w, 3% w/w, 4% w/w, 6% w/w, 8% w/w, 10%w/w, 12% w/w, 14% w/w, 16% w/w, 17% w/w, 18% w/w, 20% w/w, 22% w/w, 24%w/w, 25% w/w, 28% w/w, 30% w/w; each value represents a separateembodiment of the invention. Preferably, the dispersion concentration isup to 17% w/w; more preferably, the dispersion concentration is 17% w/w.

For use in fiber coating methods, the SP1/CB dispersion is normallydiluted to a concentration of between 0.01% and 0.5% w/w. Preferably,the concentration is between 0.05% and 0.15% w/w. More preferably theconcentration is 0.05%, 0.1% or between 0.05% and 0.1% w/w; each valuerepresents a separate embodiment of the invention.

The SP1/CB dispersions can be formed in various CB:SP1 ratios as stabledispersion. In one embodiment, the CB:SP1 ratio in the dispersion ratiois between 0.1:1 to 15:1 dry weight/weight. Preferably the CB:SP1 ratiois between 0.3:1 to 10:1 dry w/w. More preferably, the CB:SP1 ratio isbetween 3:1 to 8:1 dry w/w. In one embodiment, the CB:SP1 ratio is 5.5dry w/w. In another embodiment, the CB:SP1 ration is 3.0 dry w/w, 3.5dry w/w, 4.0 dry w/w, 4.5 dry w/w, 5.0 dry w/w, 5.5 dry w/w, 6.0 dryw/w, 6.5 dry w/w, 7.0 dry w/w, 7.5 dry w/w, 8.0 dry w/w, 9.0 dry w/w, or10.0 dry w/w; each value represents a separate embodiment of theinvention. In one embodiment the SP1 is an isolated SP1. In anotherembodiment, the SP1 is the crude SP1, i.e. the heat stable fraction ofan extract of bacteria expressing the SP1 protein.

In one embodiment, this invention is directed to a polymer, fiber, film,fabric or polymeric fabric coated with at least one layer of acomposition of matter comprising SP1 based polypeptide, CB (SP1/CB) andas described herein above. In another embodiment, the polymer, fiber,film, fabric or polymeric fabric comprises at least one of: cotton,wool, silk, nylon, polyester, polypropylene, glassfiber, elastane,Kevlar and aramid. In another embodiment, the polymer, fiber, film,fabric or polymeric fabric is polyester. In another embodiment, thepolymer, fiber, film, fabric or polymeric fabric is Aramid. In anotherembodiment, the polymer, fiber, film, fabric or polymeric fabric isnylon. In another embodiment, the nylon is a nylon fabric. In anotherembodiment the nylon is a nylon film.

In another embodiment, the polymer, fiber, film, fabric or polymericfabric according to this invention further comprises at least one layerof polyethyleneimine (PEI). In another embodiment, the polymer, fiber,film, fabric or polymeric fabric according to this invention is coatedwith at least one layer of polyethyleneimine (PEI). In anotherembodiment, the PEI is high molecular weight PEI. In another embodimentthe PEI is low molecular weight PEI. In one embodiment, the PEI thatfinds utility in the context of this invention has a molecular weight ofbetween 800 Da and 750,000 Da. In another embodiment, the PEI is highmolecular weight PEI. In another embodiment the PEI is low molecularweight PEI. In another embodiment, the molecular weight of the PEI isbetween 1 KDa and 10,000 KDa; In another embodiment, between 10 KDA and100 KDa; In another embodiment, between 25 KDa and 80 KDa; In anotherembodiment, between 50 KDa and 70 KDa. In another embodiment, themolecular weight of the PEI is about 60 KDa. In another embodiment, theapplied loading of the PEI on the fiber, film or fabric is between about0.05% and about 5%. In another embodiment, between about 0.5% and about2.5%. In another embodiment, between about 0.5% and about 1.5%. Inanother embodiment, between about 1.8% and about 2.0%. In anotherembodiment, between about 0.05% and about 0.3%. In another embodiment,the applied loading of the PEI on the fiber, film or fabric is 0.05%,0.07%, 0.09%, 0.1%, 0.12%, 0.14%, 0.16%, 0.18%, 0.2%, 0.21%, 0.25%,0.28%, or 0.3%; each value represents a separate embodiment of theinvention.

In another embodiment, the loading of said composition on said polymer,fiber, film, fabric or polymeric fabric is between 5 gr/kg and 15 gr/kg.In another embodiment, the loading is between 7 gr/kg and 14 gr/kg. Inanother embodiment, the loading is about 7 gr/kg. In another embodiment,the loading is about 10 gr/kg. In another embodiment, the loading isabout 10.5 gr/kg. In another embodiment, the loading is about 14 gr/kg.In another embodiment, the loading is about 3.5 gr/kg. In anotherembodiment, the loading is about 1 gr/Kg, 2 gr/Kg, 3 gr/Kg, 3.5 gr/Kg, 4gr/Kg, 5 gr/Kg, 5.5 gr/Kg, 6 gr/Kg, 7 gr/Kg, 8 gr/Kg, 9 gr/Kg, 10 gr/Kg,11 gr/Kg, 12 gr/Kg, 14 gr/Kg, 16 gr/Kg, 18 gr/Kg, 20 gr/Kg, 30 gr/Kg, 40gr/Kg, 50 gr/Kg, 60 gr/Kg, 70 gr/Kg, 80 gr/Kg, 90 gr/Kg, 100 gr/Kg; eachvalue represents a separate embodiment of the invention. Preferably, theloading is 7 gr/Kg. The loading can be achieved by one layer coating orby multi-layer coating; in one embodiment, the loading is achieved byone layer coating; in another embodiment, the loading is achieved by twolayer coating; in another embodiment, the loading is achieved by threelayer coating; preferably the coating is achieved by two layer coating;e.g. two loadings of 3.5 g/Kg.

In one embodiment, this invention is directed to a polymer, fiber, film,fabric or polymeric fabric coated with a composition of mattercomprising SP1 based polypeptide and CB according to this invention(i.e., SP1/CB composition).

In one specific embodiment, SP1 based polypeptides and chimeric SP1polypeptides, are used to bind carbon black, and the resulting SP1polypeptide-CB-complex (i.e., SP1/CB composition) is then used to bindthe CB to a polymer fiber, film or fabric, such as aramid (e.g.Kevlar™), for incorporation into rubber tires, in order to enhance themechanical and/or physical properties (e.g. electrical, mechanicaland/or thermal) and function of the tires.

As used herein, the phrase “SP1 based polypeptide-CB complex” or “SP1/CBcomposition” refers to a composition comprising an SP1 or chimeric SP1polypeptide, bound to carbon black.

As used herein, the phrase “SP1 polypeptide-CB-polymer complex” orSP1/CB-enforced fiber” refers to a polymer, fiber, yarn, cord, film orfabric coated with an SP1/CB composition according to this invention.

This invention also relate to methods of producing fibers, yarns, cords,films, fabrics or polymeric fabrics coated with carbon nanoparticles(e.g., carbon black or carbon nanotubes). In one embodiment, the methodfor producing such fibers, yarns, cords, films, fabrics or polymericfabrics comprise contacting a dispersion comprising composition ofmatter comprising SP1 based polypeptide according to this invention, CB(SP1/CB) with a polymer, fiber, film, fabric or polymeric fabric.Preferably, the method comprises a prestep of de-sizing of said fibers,yarns, cords, films, fabrics or polymeric fabrics prior to contacting itwith the composition. In one embodiment, the method further comprises astep of washing the unbound composition from said polymer, fiber, film,fabric or polymeric fabric. In another embodiment, the method furthercomprises a step of contacting the polymer, fiber, film, fabric orpolymeric fabric, with polyethyleneimine (PEI) prior to contacting thepolymer, fiber, film, fabric or polymeric fabric with said composition,and after the de-sizing pre-step. Preferably, the steps of contactingthe dispersion with the fiber, washing the unbound composition andcontacting with PEI are repeated at least once. In one embodiment, themethod further comprises a post-treatment step of contacting the fiber,yarn, cord, film, fabric or polymeric fabric with PEI as the last step.In another embodiment, said step of contacting the dispersion comprisingthe composition of this invention with the polymer, fiber, film, orfabric, is performed using a textile dying machine. Non limitingexamples of textile dying machines are: Jigger coating machine for wovenfabrics or vertical or horizontal yarn or fabric package dyeing system.

In one embodiment, this invention is directed to a method of producing afiber, yarn, cord, film, fabric or polymeric fabric coated with carbonblack, said method comprises:

-   -   a. Optionally de-sizing a fiber, yarn, cord, film, fabric or        polymeric fabric;    -   b. Optionally contacting said fiber, yarn, cord, film, fabric or        polymeric fabric with polyethyleneimine (PEI);    -   c. Contacting a dispersion comprising a composition of SP1 Based        polypeptide, CB with the fiber, yarn, cord, film, fabric or        polymeric fabric;    -   d. Optionally repeating steps (b) and (c) at least once;    -   e. Optionally contacting said fiber, yarn, cord, film, fabric or        polymeric fabric, with polyethyleneimine (PEI).

In another embodiment, said step of contacting the dispersion comprisingthe composition of this invention with the polymer, fiber, film, orfabric, is performed using a textile dying machine.

Methods of preparing composite materials using SP1/CB complexes include,but are not limited to contacting an SP1/CB dispersion in a solvent withpolymer, fiber, film, or fabric under conditions sufficient to form anSP1/CB-polymer composite, wherein the SP1/CB complex is deposited on thepolymer, fiber, film or fabric. In one embodiment, the polymer, fiber,film or fabric is dipped into the SP1/CB dispersion to form a SP1/CBcoated composite material. The solvent used for the SP1/CB dispersioncan be water, common organic solvents or a mixture thereof.

In one specific embodiment, SP1 based polypeptides and chimeric SP1polypeptides, are used to hind carbon nanotubes, and the resulting SP1polypeptide-CNT-complex is then used to bind the CNT to a polymer fiber,film or fabric, such as aramid (e.g. Kevlar™).

As used herein, the phrase “SP1 based polypeptide-CNT complex” or“SP1/CNT” refers to a composition comprising an SP1 or chimeric SP1polypeptide, bound to at least one carbon nanotube, for example, asdescribed in detail in Examples 1-3. This invention also relate tomethods of producing fibers, yarns, cords, films, fabrics or polymericfabrics coated with carbon nanotubes. In one embodiment, the method forproducing such fibers, yarns, cords, films, fabrics or polymeric fabricscomprise contacting a dispersion comprising composition of mattercomprising SP1 based polypeptide according to this invention, CNT with apolymer, fiber, film, fabric or polymeric fabric. Preferably, the methodcomprises a pre-step of de-sizing of said fibers, yarns, cords, films,fabrics or polymeric fabrics prior to contacting it with thecomposition. In one embodiment, the method further comprises a step ofwashing the unbound composition from said polymer, fiber, film, fabricor polymeric fabric with buffer or water. In another embodiment, themethod further comprises a step of contacting the polymer, fiber, film,fabric or polymeric fabric, with polyethyleneimine (PEI) prior tocontacting the polymer, fiber, film, fabric or polymeric fabric withsaid composition, and after the de-sizing prestep. Preferably, the stepsof contacting the dispersion with the fiber, washing the unboundcomposition and contacting with PEI are repeated at least once. In oneembodiment, the method further comprises a post-treatment step ofcontacting the fiber, yarn, cord, film, fabric or polymeric fabric withPEI as the last step. In another embodiment, said step of contacting thedispersion comprising the composition of this invention with thepolymer, fiber, film, or fabric, is performed using a textile dyingmachine. Non limiting examples of textile dying machines are: Jiggercoating machine for woven fabrics or vertical or horizontal yarn orfabric package dyeing system.

In one embodiment, a method of producing a fiber, yarn, cord, film,fabric or polymeric fabric coated with carbon nanotubes, includes:

-   -   a. Optionally de-sizing a fiber, yarn, cord, film, fabric or        polymeric fabric;    -   b. Optionally contacting said fiber, yarn, cord, film, fabric or        polymeric fabric with polyethyleneimine (PEI);    -   c. Contacting a dispersion comprising a composition of SP based        polypeptide, CNT according to this invention with the fiber,        yarn, cord, film, fabric or polymeric fabric;    -   d. Optionally repeating steps (b) and (c) at least once; and    -   e. Optionally contacting said fiber, yarn, cord, film, fabric or        polymeric fabric, with polyethyleneimine (PEI).

In a certain embodiment, washing is performed at each step with abuffer, water, or other appropriate scouring agent.

Contacting the dispersion with the polymer, fiber, film, or fabric, isperformed either in a textile dying machine for batch processing or in abath for continuous processing depending on the embodiment.

Various techniques are suitable for the formation of nanocompositematerials. These include injection molding, extrusion, blow molding,thermoforming, rotational molding, cast and encapsulation andcalendaring.

Definitions

As used herein, a plurality of filaments or fibers grouped togethereither through twisting or entanglement forms a yarn strand alsoreferred to as yarn. A plurality of yarn wound together forms thread anda plurality of threads wound together forms cord. Yarn, thread, or cordcombined into a sheet either through weaving or otherwise forms fabric.

A “fiber” refers to a strand having a diameter between 1-1000 micron.

“Tex” unit refers to mass in grams of fiber/1,000 meters and a decitex(dtex) refers to grams of fiber/10,000 meters.

“High twist” refers to more than 120 twists of yarn or thread/meter andlow twist refers to at less than 80 twists of yarn or thread/meter.Medium twist refers to 80-120 twists/meter. Twist levels applies tofibers, yarn, and thread.

“Interlocked fibers” refers to state in which a plurality of fibers areheld together either through entanglement or twisting with each other,according to an embodiment.

“Coating” is defined by the type material forming the coating; a coatingthat is formed from material different from the adjacent material isdeemed as a separate coating. For example, three coatings in which amaterial differing from the outer coatings would constitutes threecoatings even if the outer layers are the same material since coatingeach is different from the adjacent material. It should be appreciatedthat each coating is formed by one or more contiguous layers.

“Room Temperature” refers to a temperature anywhere between 20°-60° C.In a certain embodiment certain steps or all steps can be performed at atemperature as low as 4° C. and as high as 90° C.

It should be appreciated that the number twists is defined by theoutmost unit and not the subunits forming the unit for the purposes ofthis application. For example, high twist thread refers to the number ofwinds of thread and not the number of winds of yarn forming the thread.Similarly, low twist yarn refers to the number of winds of yarn and notthe number of winds of the fibers forming the yarn.

As used herein the term “about” refers to ±10%.

Throughout this application, various embodiments of this invention maybe presented in a range format. It should be understood that thedescription in range format is merely for convenience and brevity andshould not be construed as a limitation on the scope of the invention.Accordingly, the description of a range should be considered to havespecifically disclosed all the possible subranges as well as individualnumerical values within that range. For example, description of a rangesuch as from 1 to 6 should be considered to have specifically disclosedsubranges such as from 1 to 3, from 1 to 4, from 1 to 5, from 2 to 4,from 2 to 6, from 3 to 6 etc., as well as individual numbers within thatrange, for example, 1, 2, 3, 4, 5, and 6. This applies regardless of thebreadth of the range.

Whenever a numerical range is indicated herein, it is meant to includeany cited numeral (fractional or integral) within the indicated range.The phrases “ranging/ranges between” a first indicate number and asecond indicate number and “ranging/ranges from” a first indicate number“to” a second indicate number are used herein interchangeably and aremeant to include the first and second indicated numbers and all thefractional and integral numerals there between.

As used herein the term “method” refers to manners, means, techniquesand procedures for accomplishing a given task including, but not limitedto, those manners, means, techniques and procedures either known to, orreadily developed from known manners, means, techniques and proceduresby practitioners of the chemical, pharmacological, biological,biochemical and medical arts.

It is appreciated that certain features of the invention, which are, forclarity, described in the context of separate embodiments, may also beprovided in combination in a single embodiment. Conversely, variousfeatures of the invention, which are, for brevity, described in thecontext of a single embodiment, may also be provided separately or inany suitable subcombination or as suitable in any other describedembodiment of the invention. Certain features described in the contextof various embodiments are not to be considered essential features ofthose embodiments, unless the embodiment is inoperative without thoseelements.

Various embodiments and aspects of the present invention as delineatedhereinabove and as claimed in the claims section below find experimentalsupport in the following examples.

Additional objects, advantages, and novel features of the presentinvention will become apparent to one ordinarily skilled in the art uponexamination of the following examples, which are not intended to belimiting. Additionally, each of the various embodiments and aspects ofthe present invention as delineated hereinabove and as claimed in theclaims section below finds experimental support in the followingexamples.

EXAMPLES

Reference is now made to the following examples, which together with theabove description, illustrate some embodiments of the invention in anon-limiting fashion.

General Experimental Concept of Complexing SP1 and CNP

The studies presented below demonstrate two strategies for altering thebinding properties of SP1 variants, namely the affinity and avidity ofSP1 variants to various substrates and controlling the immobilization ofSP1 on various surfaces. Affinity and avidity are two terms used inprotein biochemistry to describe strength of non-covalent interactions,the phenomenon whereby certain atoms or molecules have the tendency toaggregate or bond.

The term “affinity” is used to describe the strength of a single bond,while the term “avidity” is use to describe the combined strength ofmultiple bond interactions affinity. Dissociation constant (Kd), orequilibrium constant, is the inverse of the affinity constant, measuresthe propensity of a larger object to separate (dissociate) reversiblyinto smaller components, as when a complex falls apart into itscomponent molecules, or when a salt dissociates into its component ions.The dissociation constant is usually denoted Kd and is the inverse ofthe affinity constant. In the special case of salts, the dissociationconstant can also be called the ionization constant.

The first strategy involves positioning of the anchoring side-chains,such as found in cysteine residues, on the dodecameric protein's ringrim, and comparing the binding properties of the resulting constructwith those of a protein construct having anchoring side-chainspositioned at the inner side of the annulus (the pore or “hole” of thering). This strategy uncovers the capacity of the SP1 basic architectureto protect certain regions on its surface, and ligands attached thereat,from surface exposure.

In the second strategy, several binding moieties are attached to the SP1dodecameric protein at the protein's annulus inner pore by geneticengineering. By fusing these specific affinity peptides at a putativeprotected part of the protein, the binding moieties are expected to beless available for binding with large entities which are excluded fromthe protein's pore. This experimental strategy is designed to study theeffect of changing the conditions of the media of the protein, and toshow that entering a factor to the media, which can affect the structureof the SP1 monomers and thus the structure of the entire dodecamer, cancontrol the degree of exposure of the binding moieties to the media. Theevent of adding the structure altering factor, such as a denaturatingagent, can thus increase the ability of the binding moieties to interactand bind large entities in the media. The capacity to switch from anon-binding entity to a binding entity by adding and removing a chemicalfactor constitutes a chemical switch.

The concept of a chemical switch was demonstrated by fusing severalspecific affinity peptides, such as silicon binding peptides, to each ofthe SP1 basic skeleton, at inner pore position, to thereby obtain asilicon binding protein switch, which is sensitive to the media levelsof denaturating agents, such as guanidinium hydrochloride (GuHCl). Theaffinity peptide was isolated by Sano and coworkers [Sano, K. I. et al.JACS. 125, pp 14234-14235, 2003; Sano, K. I et al., JACS, 128, pp1717-1722, 2006; and Sano, K. I. et al., Nano Lett., 7, pp 3200-3202,2007] using a peptide-phage display system. This six amino acidspeptide, referred to herein and in the art as mTBP, was reported by Sanoand coworkers to bind to Ti, Ag and Si surfaces, but not to Au, Cr, Pt,Sn, Zn, Cu, or Fe.

Thus, a SP1 scaffold was modified to present 12 copies of the mTBPhexapeptide in a switchable manner. A positive cooperative effect isdemonstrated when the peptide is presented on the SP1 dodecamer, ascompared to the free peptide, accompanied with significant reduction innon-specific binding of the fused peptides compared to that of the freepeptide.

Example 1 Construction of SP1 Variants with High Affinity to VariousMaterials

WO 2007/007325 provides a non-limiting list of peptides formingcomplexes with inorganic ionic substances, adapted from Sarikaya et al.[Ann. Rev. Mater. Res., 2004, 34, 373-408]. These relatively shortpeptides are suitable for fusion to the SP1 protein as part of themodification of the SP1 polypeptide. Many more examples of peptides withhigh affinity to different materials are disclosed in the literature.

Table 1 presents the SP1 variants used in this context, their bindingability, primers used for their construction, mutation or insertion atthe N-terminus, SP1 template, reference, and growthconditions/induction. All mutant proteins demonstrated characteristicssimilar to the wild type SP1 in terms of heat stability, proteaseresistance and complex formation. Standard nomenclature of mutations isused i.e., amino acids position using wild type sequence including firstmethionine residue.

TABLE 1 Mutation SP1 variant/ and/or Growth Relevant Insertion atSP1 Template conditions/ activity PCT Primers the N-terminusand reference induction Wild type U.S. patent Terrrfic broth or SP1application No. Luria broth/ (SEQ ID 2006/0172298 37° C./ NO: 4)IPTG 1 mM Δ2-6 Δ2-6 Wang et al. Luria broth/ (SEQ ID (2006); 37° C./NO: 64) WO IPTG 1 mM 2007/007325 M43C Δ2-6 5′ CTGCTCGAT M43C Δ2-6 Δ2-6Luria broth/ (SEQ ID CTCATTCCAAGC 37° C./ NO: 1) TGTAAGAGTTTC IPTG 1 mMAATTGGGGCAC G 3′ L81C Δ2-6 5′ GCAAGTCTG L81C Δ2-6 Δ2-6 Luria broth/Flat gold GTTTGCAAGAGT 37° C./ binding ACTGCGATTCTG IPTG 1 mM (SEQ IDCTGCTCTTGCT NO: 2) G 3′. mtbSP 5′-AAAACATAT RKLPDAA M43C Δ2-6Terrific broth or Switchable GCGCAAACTTCC (SEQ ID NO: 5) Medalsy etLuria broth/ silicon oxide GGATGCGGCAAC al. (2008); 37° C./ bindingCAGAACTCCAAA WO IPTG 1 mM CNT GCTTG-3′ and 2007/007325 dispersion SP1rev(SEQ ID 5′-AAAAGAGCT NO: 3) CTTAGTAAAGAA AGTAATCAATAA C-3′) L1-SP15′AAGGAGATAT HWSAWWIRSNQS Wild type Terrific broth/ CNT ACAAAAACATAT(SEQ ID NO: 10) 28° C./ dispersion GCACTGGTCAGC IPTG 1 mM (SEQ IDATGGTGGATACG NO: 6) ATCAAATCAATC AGCAACCAGAAC TCCAAAG 3′ 5′CTTTGGAGTTCTGGTTGCTGAT TGATTTGATCGT ATCCACCATGCT GACCAGTGCATA TGTTTTTGTATATCTCCTT 3′ L2-SP1 5′AGAAGGAGAT HSSYWYAFNNKT Wild type Terrific broth/CNT ATACAAAAACAT (SEQ ID NO: 11) 37° C./, dispersion ATGCACTCATCAIPTG 0.1 mM (SEQ ID TACTGGTACGCA NO: 14) TTCAACAACAAA ACAGCAACCAGAACTCCAAAGC 3′ 5′GCTTTGGAGT TCTGGTTGCTGT TTTGTTGTTGAA TGCGTACCAGTATGATGAGTGCAT ATGTTTTTGTAT ATCTCCTTCT 3′ L3-SP1 5′ATACAAAAAC DYFSSPYYEQLFWild type Terrific broth/ CNT ATATGGATTATT (SEQ ID NO: 12) 37° C./dispersion TTTCATCACCAT IPTG 0.5 mM Kevlar ATTATGAACAAT bindingTATTTGCAACCA (SEQ ID GAACTCC 3′ NO: 8) 5′GGAGTTCTGG TTGCAAATAATTGTTCATAATATG GTGATGAAAAAT AATCCATATGTT TTTGTAT 3 L6-SP1 5′AGAAGGAGATSNQS Wild type IPTG 1 mM/ CNT ATACAAAAACAT (SEQ ID NO: 13) 37° C./dispersion ATGTCAAATCAA Terrific broth (SEQ ID TCAGCAACCAGA NO: 16)ACTCCAAAGC 3′ 5′GCTTTGGAGT TCTGGTTGCTGA TTGATTTGACAT ATGTTTTTGTATATCTCCTTCT 3

Example 2 Carbon Nanotubes (CNT) Dispersion by SP1 Variants

The Examples presented below provide SP1 variants, fused to CNT-bindingpeptides, which are capable of binding to CNT and thereby enable theaqueous dispersion of these protein-coated CNT. Several examples ofshort peptides that were isolated from phage display libraries asCNT-binding peptides are disclosed in the literature. See, for example,Nature materials, 2003, 2, 196; Nano lett., 2006, 6, 40-44; andLangmuir, 2004, 20, 8939-8941).

Table 2 below presents the terminus sequence of these variants, as wellas their purification method and grade, N-terminal sensitivity todigestion by alcalase, and the SP1 variant concentration which isrequired for CNT dispersion. All mutant proteins demonstratedcharacteristics similar to the wild type SP1 in terms of heat stability,protease resistance and complex formation. Shift in molecular weightrelatively to samples that were not treated with alcalase was observedboth in samples that were not boiled or boiled in SDS gel applicationbuffer (complex and monomer, respectively). In all cases the apparentmolecular weight of the alcalase treated SP1 variants was higher thanthose of wild type, indicating that some but not all the addedamino-acids were removed, and they are different from publishedsequences.

TABLE 2 SDS PAGE analysis N-terminal SP1 concentration sensitivityrequired for CNT Peptide fused to Complex to digestion dispersion SP1variant the N-terminus Grade Formed by alcalase (mg/ml) References Wildtype None 80° C. plus alcalase Yes No 1 treatment Ion exchange purifiedYes <1 protein mtbSP RKLPDAA 80° C. treatment Yes No 0.2 U.S.Application No. (SEQ ID NO: 5) Ion exchange purified Yes 0.1 20070112174protein U.S. Application No. 20070117148 Nano Lett., 2007, 6, 1579-1579.L1-SP1 HWSAWWIRSNQS 80° C. plus alcalase Yes Yes 0.004 U.S. Pat. No.7,304,128 (SEQ ID NO: 10) treatment U.S. Application No. Ion exchangepurified Yes 0.004 20070117147 Ion exchange purified Yes 0.004 U.S.Application No. plus alcalase treatment 20070117150 U.S. Application No.20070117148 U.S. Application No. 20040058457 Nature Materials, 2003, 2,196 L2-SP1 HSSYWYAFNNKT 80° C. plus alcalase Yes Yes 0.04 U.S.Application No. (SEQ ID NO: 11) treatment 20060172282 Dissolvedinclusion No Complete 0.100 Nano Lett., 2006, 6, 40-44 bodies digestionRefolding of IB s Yes Yes 0.1 L3-SP1 DYFSSPYYEQLF Refolding of IBs 80°C. Yes Small shift 0.1 U.S. Application No. (SEQ ID NO: 12) plusalcalase treatment 20050277160 80° C. plus alcalase Yes Small shift 0.01Langmuir, 2004, 20, 8939-8941 treatment L6-SP1 SNQS 80° C. plus alcalaseYes No 0.05 (SEQ ID NO: 13) treatment

Surprisingly, treatment with alcalase and partial digestion of theN-terminus doesn't reduce its ability to disperse CNT. This is probablybecause in each complex not all N-termini are digested and the L1variant (SEQ ID NO: 6) complex appears as a double band. For example,N-terminus sequencing and MALDY-TOF analysis of alcalase treated L1-SP1revealed that 8 amino acids were digested by the protease and theN-terminus was SNQS but the digestion doesn't reduce its ability todisperse CNT. In agreement with this conclusion insertion of the SNQSpeptide to SP1 N-terminus yields a variant, L6 (SEQ ID NO: 15), withlower CNT dispersion activity, lower than L1 (SEQ ID NO: 6) (50-100μg/ml versus 4 μg ml, respectively).

Example 3 Tri-Complexes of SP1 Variants, CNT and Aramid (KEVLAR™) orEpoxy Resin

The capacity of the SP1 variants of the present embodiments to bind toadvanced materials, such as KEVLAR™, was studied and demonstrated, aspresented in detail in U.S. Pat. No. 8,957,189 herein incorporated byreference in its entirely.

SP1 variants were studied for their capacity as multi-functionalreagents which can bind CNT through the N-terminus to form a SP1/CNTcomplex, which in turn can bind to epoxy resin through exposed primaryamines. Such reagents can be highly useful in many practicalapplications involving water interfaces with CNT, including dispersionin epoxy resin. While water and other mediating solvents can be removedby combination of ultra-filtration and freeze drying, these processesare energy consuming and hard to control. The process presented hereintakes advantage of the fact that SP1 precipitates in the presence of70-80% ethanol after 2 hours incubation at −20° C., and so does the CNTprotein complex. The precipitated SP1/CNT complex can be easilydispersed in water. The freeze-dried precipitated SP1/CNT complex can bedispersed in epoxy resin as demonstrated in U.S. Pat. No. 8,957,189herein incorporated by reference in its entirely.

In addition, it was hypothesized that if CNT and aramid (e.g. KEVLAR™—abrand name of a strong and heat-resistant aramid fiber developed byDuPont and used in bullet-proof vests, tires, fiber-optic cables andmore) bind to SP1 variants in a similar fashion, the protein may serveas an adhesive mediator to promote attachment of these two components toeach other, based on the two-sided doughnut shape of SP1 which exhibitsbinding sites on both sides of the annulus.

Materials and Methods Bacterial Strain and Culture Conditions

Escherichia coli strain DH5α was used for cloning and E. coli strainBL21 (DE3) was used for expression. Cells were grown in either LuriaBertani medium (ΔNSP1, M43CΔNSP1 and L81CΔNSP1), Terrific broth (L1-SP1,L2-SP1, L3-SP1, L6-SP1), or either Luria or Terrific brothinterchangeably (native SP1, mtbSP), at 37° C. (except for L1-SP1, whichwas grown at 28° C.). After induction with isopropylP-D-thiogalactopyranoside (IPTG)(1 mM for native SP1, mtbSP1, ΔNSP1,M43CΔNSP1, L81CΔNSP1, L1-SP1 and L-6 SP1, 0.5 mM for L3-SP1 and 0.1 mMfor L2-SP1) bacteria were grown for additional 4 hours, followed byharvesting by centrifugation at 14,000×g for 15 minutes.

Vector Construction

Both M43C ΔNSP1 mutant and L81C ΔNSP1 mutant were constructed using sitedirected mutagenesis on the ΔNSP1 coding sequence (SEQ ID NO: 7)template (previously described by Medalsy et al. [Nano lett., 8,473-477, 2008]), performed in accordance to the Stratagene Quickchange(Stratagene, La Jolla, Calif.) protocol with the PfuTurbo or Deep-ventDNA polymerase.

Primers used for site directed mutagenesis were: C43 (5′CTGCTCGATCTCATTCCAAGCTGTA AGAGTTTCAATTGGGGCACG 3′) (SEQ ID NO: 65) forM43C, and C81 (5′ GCAAGTCTGGTTTGCAAGA GTACTGCGATTCTGCTGCTCTTGCTG 3′)(SEQ ID NO: 66) for L81C.

The mtbSP1 mutant (SEQ ID NO: 3) was constructed using 2 primers: mTBforward primer (5′AAAACATATGCGCAAACTTCCGGATGCGGCAACCAGAACTCCAAAGCTG-3′)(SEQ ID NO: 67) and SP1 reverse primer(5′-AAAAGAGCTCTTAGTAAAGAAAGTAATCAATAAC-3′) (SEQ ID NO: 68) with M43CΔNSP1 coding sequence (SEQ ID NO: 69) as a template.

The L1-SP1CNT mutant (SEQ ID NO: 6) was constructed using the primers:forward primer (5′AAGGAGATATACAAAAACATATGCACTGGTCAGCATGGTGGATACGATCAAATCAATCAGCAACCAGAACTCCAAAG 3′) (SEQ ID NO: 70) and reverse primer(5′CTTTGGAGTTCTGGTGCTGATTGATTTGATCGTATCCACCATGCTGACCAGTGCATATGTTTTTGTATATCTCCTT 3′) (SEQ ID NO: 71) with native SP1 codingsequence (SEQ ID NO: 28) as a template.

The L2-SP1CNT mutant (SEQ ID NO: 14) was constructed using the primers:forward primer (5′AGAAGGAGATATACAAAAACATATGCACTCATCATACTGCGTACGCATTCAACAACAAAACAGCAACCAGAACTCCAAAGC 3′) (SEQ ID NO: 72) and reverse primer(5′GCTTTGGAGTTCTGGTTGCTGTTTTGTTGTTGAATGCGTACCAGTATGATGAGTGCATATGTTTTTGTATATCTCCTTCT 3′) (SEQ ID NO: 73) with native SP1 codingsequence (SEQ ID NO: 28) as a template.

The L3-SP1CNT mutant (SEQ ID NO: 12) was constructed using the primers:forward primer (5′ATACAAAAACATATGGATTATTTTTCATCACCATATTATGAACAATTATTTGCAACCAGAACTCC 3′) (SEQ ID NO: 74) and reverse primer(5′GGAGTTCTGGTTGCAAATAATTGTTCATAATATGGTGATGAAAAATAATCC ATATGTTTTTGTAT) 3(SEQ ID NO: 75) with native SP1 coding sequence (SEQ ID NO: 28) as atemplate.

The L6-SP1CNT mutant (SEQ ID NO:15) was constructed using the primers:forward primer (5′AGAAGGAGATATACAAAAACATATGTCAAATCAATCAGCAACCAGAACTCCAAAGC 3′) (SEQ ID NO: 76) and reverse primer (5′GCTTTGGAGTTCTGGTTGCTGATTGATTTGACATATGTTTTTGTATATCTCCTT CT 3) (SEQ ID NO:77) with native SP1 coding sequence (SEQ ID NO: 28) as a template.

The L7-SP1CNT mutant (SEQ ID NO: 16) is identical to L1-SP1CNT sequence,except for mutation of the nucleotide sequence encoding the insertedpeptide at 5Ile from ata to att, and at 6Arg from cga to cgt, to improvecodon usage. The mutant polypeptide was constructed using the primers:for A24T mutant, forward primer(5′-ACTGGTCAGCATGGTGGATTCGATCAAATCAATCAG-3′) (SEQ ID NO: 78) and reverseprimer (5′-CTGATTGATTTGATCGAATCCACCATGCTGACCAGT-3′) (SEQ ID NO: 79). ForA27T mutant, forward primer(5′-GTCAGCATGGTGGATTCGTTCAAATCAATCAGCAACC-3′) (SEQ ID NO: 80) andreverse primer (5′-GGTTGCTGATTGATTTGAACGAATCCACCATGCTGAC-3′) (SEQ ID NO:81), using “QuikChange Site-Directed Mutagenesis Kit” of “Stratagene”(La Jolla, Calif.).

The L4-SP1CNT mutant (SEQ ID NO: 9) is identical to L1-SP1 CNT sequence,except for mutation of R23K of the inserted peptide. The mutantpolypeptide was constructed using the primers: for R23K mutant, forwardprimer (5′-TGACTCGGTTCAAGGATGAGATCACAAAAGAACAGATCGACA-3′) (SEQ ID NO:82), and reverse primer(5′-TGTCGATCTGTTCTTTTGTGATCTCATCCTTGAACCGAGTCA-3′) (SEQ ID NO: 83) using“QuikChange Site-Directed Mutagenesis Kit” of “Stratagene” (La Jolla,Calif.).

The L5-SP1CNT mutant (SEQ ID NO: 17) is identical to L1-SP1CNT sequence,except for mutation of T22C of the inserted peptide. The mutantpolypeptide was constructed using the primers: for T22C mutant, forwardprimer (5′-ACTCGGTCAAGGATGAGATCTGCCGAGAACAGATCGACAACTAC-3′) (SEQ ID NO:84), and reverse primer(5′-GTAGTTGTCGATCTGTTCTCGGCAGATCTCATCCTTGAACCGAGT-3′) (SEQ ID NO: 85)using “QuikChange Site-Directed Mutagenesis Kit” of “Stratagene” (LaJolla, Calif.).

The L8-SP1CNT mutant (SEQ ID NO: 18) is identical to L4-SP1CNT sequence,except for mutation of the nucleotide sequence encoding the insertedpeptide at 5Ile from ata to att, and at 6Arg from cga to cgt, to improvecodon usage. The mutant polypeptide was constructed using the primers:for A24T mutant, forward primer(5′-ACTGGTCAGCATGGTGGATTCGATCAAATCAATCAG-3′) (SEQ ID NO: 78) and reverseprimer (5′-CTGATTGATTTGATCGAATCCACCATGCTGACCAGT-3′) (SEQ ID NO: 79). ForA27T mutant, forward primer(5′-GTCAGCATGGTGGATTCGTTCAAATCAATCAGCAACC-3′) (SEQ ID NO:80) and reverseprimer (5′-GGTTGCTGATTGATTTGAACGAATCCACCATGCTGAC-3′) (SEQ ID NO:81),using “QuikChange Site-Directed Mutagenesis Kit” of “Stratagene” (LaJolla, Calif.).

Wild type SP1 was used as a template for PCR reaction(5′-ACTGGTCAGCATGGTGGATTCGATCAAATCAATCAG-3′) (SEQ ID NO: 78) and reverseprimer (5′-CTGATTGATTTGATCGAATCCACCATGCTGACCAGT-3′) (SEQ ID NO: 79) withnative SP1 coding sequence (SEQ ID NO: 28) as a template.

All constructs were inserted into pET 29a expression plasmid (NovagenInc. Madison Wis., USA).

Protein Purification and Refolding

After centrifugation, cell pellets were resuspended in lysis buffer (50mM Tris HCL 1 mM EDTA, 10 mM MgCl2, pH 8) and sonicated on ice forseveral minutes with pulsed bursts. Variants were expressed as solubleproteins [mtbSP (SEQ ID NO: 3), L1-SP1 (SEQ ID NO: 6), L6-SP1 (SEQ IDNO: 15)], or aggregated into inclusion bodies [L2 SP1 (SEQ ID NO: 14)and L3-SP1 (SEQ ID NO: 8)].

The insoluble pellets were separated by centrifugation at 14000×g for 15minutes. Soluble mutated proteins (M43C ΔNSP1 and mtbSP1; L1-SP1;L2-SP1; L3-SP1; L6-SP1) were then heat treated at 85° C. for 30 minutesand treated by protease (alcalase, Novozyme 10<6>-fold dilution: 30 min40° C.)

Inclusion bodies of L81C ΔNSP1 (SEQ ID NO: 2) mutant were washed firstfor 15 minutes with the IB washing buffer (20 mM Tris HCL, 2 M urea, pH8) and thereafter centrifuged at 14000×g for 15 minutes. The pelletswere resuspended in denaturation buffer (20 mM Tris HCl, 6 M urea, 10 mMdithiothreitol, pH 8) and diluted to protein concentration of 5 mg/ml.Denaturated proteins were then refolded by dialysis against a foldingbuffer (20 mM Tris HCl, 1 mM DTT, pH 7) for 4 days.

Ion Exchange FPLC

Hitrap Q Sepharose XL column (1 ml) (Amersham Biosciences, Piscataway,N.J. USA), was used to purify the proteins. Samples were loaded on thecolumn using 20 mM piperazine pH 6.3 buffer at a flow rate of 3 ml/minElution was conducted with a gradient of 1 M NaCl in the same buffer anddetermined at 27-33% salt. (mTBP Appendage Peptide: mTBP peptide (SEQ IDNO: 5) was synthetically manufactured by BioSight ltd. (Karmiel,Israel).

Stability Characterization of Mutated Proteins

Three different stability analyses were performed on the wild-type SP1(SEQ ID NO: 4) and each of the mutated proteins.

-   -   1. Heat treatment (H.T) at 80° C. for 30 minutes;    -   2. Boiling treatment (B.T.) at 100° C. for 30 minutes; and    -   3. Resistance to proteolysis by proteinase K (PK) at a        concentration of 50 ug/ml of the enzyme for one hour at 37° C.        PK was eliminated by B.T. for 5 minutes.

Alternatively, alcalase was used to determine stability: Alcalase(Novozyme, 1:1000 dilution) was added at 40° C. for 30 mM Reaction wasstopped by inhibition of alcalase at 80° C. for 30 min.

All treatment were followed by centrifugation at 14,000×g for 15minutes, and analyzed by SDS-PAGE.

Silica Binding

mtbSP1 (SEQ ID NO: 3) was mixed with 10 mg silica gel (product no:28,860-8, Sigma-Aldrich, USA) in 10 mM MES pH 6.5, 150 mM NaCl, with orwithout 3M GuHCl. The solution was then incubated for one hour on arotary shaker at room temperature. Thereafter the silica was washedthree times with the same buffer without GuHCl. Bound protein wasanalyzed either by SDS-PAGE or by measuring protein concentration usingthe Micro BCA assay kit (Pierce, Rockford, USA).

Surface Preparation and Binding SP1/CNT Binding

SP1/CNT binding to aramid was evaluated using three methods:

-   -   1. Determination of the difference between CNT content in        solution (suspension) before and after its binding to the        fabric. CNT content of a suspension is determined by        precipitating the SP1/CNT from a sample of the suspension using        guanidinium hydrochloride (100 mM) or HCl (0.3%), before and        after its incubation with the fabric (combined with the washing        solution), drying the pelleted CNT, and weighing;    -   2. Spectroscopy: CNT content can be evaluated using        spectroscopic method, namely, light transmittance by        visualization of a fabric or surface coated by CNT at high        resolution under a scanning electron microscope (HR-SEM); and    -   3. Surface resistivity-CNT content of a coated fabric or surface        can be assessed by measuring surface resistivity to current flow        (this method is relevant only in cases in which the untreated        fabric is a insulator or very poor conductor).

Example 4 Carbon Nanotubes (CNT) Dispersion by SP1 Variants

Materials and Methods:

Multi wall carbon nanotubes (MWCNT) were obtained either from ArkemaInc., France (GRAPHISTRENGTH™ C100) or from Bayer MaterialScience AG,Germany (Baytubes C150 P). Single wall carbon nanotubes (SWCNT) wereobtained from Teijin, Ltd (Yamaguchi, Japan).

For small scale production, between 1.0 and 1.3 mg of MWNTs were weighedin 1 ml screw-cap glass tube (Fisherbrand, cat. no. 03-338 AA, size12×35 mm, ½ DR). A 1 ml protein solution in NaPi buffer (10 mM; pH 8.0)was added to the screw-cap glass tubes containing pre-weighted MWNTs.The resulting mixture was sonicated for 2 hours at 80° C. using an ElmaTranssonic Sonifier. The sonicated samples were first centrifuged in anEppendorf centrifuge tube for 20 minutes at 20000×g. Ninety percent ofthe upper supernatant was separated using a pipette, avoiding taking thesediment at the bottom, and transferred to another Eppendorf centrifugetube. The separated supernatant samples were diluted ten-fold. The CNTdispersion by L2-SP1 (SEQ ID NO: 14) was also tested in Tris buffer (10mM; pH 8.0) with or without urea.

For larger scale production, 400-mg of MWCNT were weighed into a glassflask, a protein solution (400 ml in NaPi buffer; 10 mM; pH 8.0) wasadded, and the mixture sonicated at a power setting of 260 W for 4hours, maintaining a maximal temperature of less than 50° C., using aMisonix 4000 Sonicator with a booster home, a 1 inch flat tip and atemperature control unit or a Hielcher sonicator (UIP1000 hd). In orderto obtain full dispersion of the sample, the sonicated samples werecentrifuged in an Eppendorf centrifuge tube for 20 minutes at 20000×guntil only a minor pellet was formed. After pelleting of the undispersedmaterial, the supernatant was very dark with the CNT, even after 100fold dilution in the same buffer. The last step was centrifugation ofthe suspension for 60 minutes at 7000 rpm using a Sorval SLA 3000centrifuge.

Results

Table 2 hereinabove, presents the results of the CNT dispersionexperiments. The SP1 variants described in Table 2, are heat stable andgenerally protease resistant, however, incubation with alcalase(1000-fold dilution) causes a shift in the molecular weight relative tosamples not treated with alcalase. In all cases the apparent molecularweight of the alcalase-treated SP1 variants was still higher than thoseof native SP1, indicating that some but not all the amino-acids derivedfrom the CNT binding peptides were removed.

As can be seen in Table 2, fusion of copies of these CNT bindingpeptides to SP1 N-terminus improves the SP1's ability to disperse multiwall carbon nanotubes (MWCNT) in a solvent, while native SP1 proteinaffords MWCNT dispersion only at high SP protein concentrations (approx.1 mg/ml). As can further be seen in Table 2, the fusion proteins have amuch higher CNT-binding activity. The greatest CNT dispersing capabilitywas observed with L1 (SEQ ID NO: 6), an SP1 polypeptide with theHWSAWWIRSNQS peptide (SEQ ID NO: 10) fused to the N-terminus, whichallowed MWCNT dispersion at the relatively low concentration 0.004mg/ml. Fusion of SP1 with the other CNT-specific peptides L2 (SEQ ID NO:14) and L3 (SEQ ID NO: 8) [HSSYWYAFNNKT (SEQ ID NO: 11) and DYFSSPYYEQLF(SEQ ID NO: 12) peptides respectively] also resulted in greater CNTdispersing activity than native SP1. Using the L3-SP1 the Hielchersonicator, maximal CNT concentration was 40 mg CNT/gr fabric, or 4%. IfSP1 solubility is as high as 150 mg/ml theoretically, maximal CNTconcentration, can be as high as 30%.

Variants L2-SP1 (SEQ ID NO: 14) and L3-SP1 (SEQ ID NO: 8) differ fromother SP1 variants in that they form both soluble and insoluble proteinfound in inclusion bodies (IB). The protein found in IB can be dissolvedin the presence of urea and refolds upon urea dilution. While properlyfolded L2-SP1 forms complexes and was protease resistant, the dissolvedL2-SP1 from IB did not form complexes and was protease sensitive. TheL2-SP1 and L3-SP1 from inclusion bodies can disperse CNT but with muchless efficiency than the soluble protein (Table 2).

As can be seen in Table 2 hereinabove, the minimal SP1 concentrationrequired for CNT dispersion depends on the sequence of the peptide fusedto the N-terminus.

In the case of L1-SP1 (SEQ ID NO: 6), an ion exchange purified proteinwas treated with alcalase, and the protein underwent N-terminussequencing and molecular weight determination using MOLDY-TOF. Theresults indicated that 8 amino acids were digested by the alcalase fromits N-terminus, leaving the SNQS peptide fused to the proteinN-terminus. It is noted herein that the SP1 variant with an insertion ofthe SNQS peptide at the N-terminus (L6-SP1, SEQ ID NO: 15) exhibitedsimilar characteristics, namely mobility in SDS PAGE and CNT dispersionability, as compared to the alcalase-treated L1-SP1.

In addition to the CNT specific peptides, fusion of the silicon/titaniumoxide binding peptide (RKLPDAA) (SEQ ID NO: 5), which yields the SP1variant mtbSP1 (SEQ ID NO: 3), was found to facilitate CNT dispersion atlower concentration than native SP1.

For industrial applications it is preferred to keep the proteinproduction costs as low as possible, and to make sure that otherpeptides that may exist in the crude extract of transformed cellsexpressing a recombinant protein do not interfere with the variant SP1'sCNT dispersion capability. To test this facet, crude extract obtainedfrom bacteria transformed to express L1-SP1 was exposed to combinationsof heat and protease treatments, and was then assessed for the retentionor loss of CNT dispersion activity, as presented in Table 3 below.

TABLE 3 Crude extract treatment CNTs dispersion Minimal protein AlcalaseMaximal concentration Heat treatment treatment dilution μg/ml Standardtreatment. 80° C., 30 minutes × 2 + 1:26 10 Heat treatment. 80° C., 120minutes − 1:26 9.3 Alcalase treatment with final 80° C., 30 minutes +1:26 6 inactivation of alcalase after alcalase treatment Alcalasetreatment without — + 1:26 11 final Inactivation of alcalase.

As can be seen in Table 3, the heat treatment of crude extract ofL1-SP1, up to 120 minutes at 80° C. and proteolysis used during thepreparation of the mutant, did not abolish the capacity to disperse CNT.

A direct demonstration that L1-SP1 binds to CNT and forms a complex wasobtained by comparing a suspension of a sample of L1-SP1 (SEQ ID NO: 6)with CNT (L1-SP1/CNT), a sample of the protein without CNT (L1-SP1) anda filtrate (0.22 micron filter) of these two samples before and afterboiling. Both the boiled and not boiled samples were analyzed by SDSPAGE.

The boiled L1-SP1 was detected as a band of the monomeric form and aband of the trimeric form, while the unboiled L1-SP1 appears as a highmolecular weight complex only.

A large fraction of the CNT was excluded by filtration, therefore longerthan 0.22 micron. The proportion of the SP1 trimer bands in the absenceof CNT was lower than that detected in the presence of CNT, both in thefiltrates and in the unfiltered samples. Apparently not all the proteindissociated upon mixing with the SDS Tricine sample buffer and SDS PAGEapplication.

Another indication that the L1-SP1 protein (SEQ ID NO: 6) binds to theCNT comes from the protein determination assay (Bradford protein assay)of both mtbSP-SP1/CNT suspension and the microfiltration (0.2 micron)flow-through, demonstrating that about 50% of the protein after CNTcomplexing is larger than the 0.2 micron pore size and is retained bythe filter (data not shown).

Yet further evidence for the formation of a SP1-CNT complex is seen inthe results of ethanol precipitation (Table 3): While uncomplexedprotein does not precipitate with 50% ethanol, SP1-CNT does precipitatewith 50% ethanol. CNT precipitate also with GuHCl (100 mM) and in acidicpH (by adding HCl or acetic acid), however, while GuHCl does not induceuncomplexed protein precipitation but acidic pHs does. This phenomenonis used to determine CNT concentration to fabrics.

Heat stability, protease and alkali resistance of L1-SP1-CNT complex: Toassess the stability of the complex, L1-SP1/CNT was subjected to heattreatment (100° C.; 10 minutes or 80° C.; 30 minutes) both at pH 8.0 andpH 11, or proteolysis at pH 8.0. Incubation of L1SP1/CNT samples, atdifferent pHs, were followed by high speed centrifugation (20 minutes;20000×g) and 10-fold dilution of the supernatant. The results of theheat and proteinase assays demonstrate that the SP1/CNT complex is heatstable and protease resistant, allowing economically desirable heatdrying and powdering of the complex prior to its dispersion in importantpolymeric compounds, such as epoxy.

The high durability of the SP1/CNT complex allowed the development ofsimple method to obtain a dry pellet of SP1/CNT complex that can easilyre-dispersed in water. The process includes first dispersion of 4% CNT,followed by three steps of wash and precipitation by 1:5 dilution inethanol (final 99% ethanol), and dehydration using a vacuum pump.

Example 5 SP1 Variants Binding to Aramid (e.g. KEVLAR™)

Material scientists and engineers are excited by the possibilities forcreating super-strong, high-performance polymer composite materialsusing carbon nanotubes. Currently, all existing methods of fabricatingCNT-polymer composites involve complicated, expensive, time-demandingprocessing techniques such as solution casting, melting, molding,extrusion, and in situ polymerization, requiring that the nanotubeseither be incorporated into a polymer solution, molten polymer or mixedwith the initial monomer before the formation of the final product (e.g.yarn, ribbon or film). This is unsuitable for insoluble or temperaturesensitive polymers, which decompose without melting.

Aramid polymers (e.g. KEVLAR™) is a well-known high-strength polymerwith a variety of important applications such as pneumatic tire treadand sidewalls, bullet-proof vests and car armor plating. However, aramid(e.g. KEVLAR™) is not soluble in any common solvent and, having nomelting point, decomposes above 400° C. As a result, aramid (e.g.KEVLAR™) fibers must be produced by wet spinning from sulphuric acidsolutions. Binding of SP1/CNT complex to aramid (KEVLAR™) was assessedfor effective post-processing incorporation of carbon nanotubes intoalready formed polymer products, such as, for example, aramid (KEVLAR™)yarns.

CNT binding to the fabric via the protein increases its surface area,allowing better interaction with the fiber and induces cross linkingbetween the fibers. In addition, protein biding to the fiber by itselfmay improve the interaction with the polymer through reactive groups onthe protein surface. It is demonstrated that some SP1 variants that bindCNT also bind to structural fibers.

Materials and Methods

L3SP1/CNT solution (SEQ ID NO: 8), in different concentrations (22μg/ml, 44 μg/ml, and 88 μg/ml samples in 10 mM NaPi, pH-8) was incubatedwith 100 mg of aramid (KEVLAR™) fabric in a rotary shaker at 25° C. for16 hours, followed by extensive wash with the same buffer to removetraces of the unbound protein and CNT, until the solution was colorless,indicating absence of CNT, and until no protein was detected in thewash. CNT binding to the aramid (KEVLAR™) was assessed by darkening ofthe aramid (KEVLAR™) fibers. SP1 binding to the washed aramid (KEVLAR™)was determined by reacting the aramid (KEVLAR™) with 2 ml of BCA proteinassay reagent (Pierce, cat No. 23227) for 30 minutes at 37° C., andmeasurement of optical density at 562 nm. The amount of protein boundwas calculated and plotted, and the results are presented in U.S. Pat.No. 8,957,189 herein incorporated by reference in its entirely.

SP1/CNT binding to aramid was evaluated by precipitation, lighttransmittance (spectroscopy, visual inspection) and surface resistivity,as detailed above.

Results

Comparison of the bound and unbound fibers after incubation with the L3SP1/CNT complex, indicated extensive binding of the CNT, even afterexhaustive washing (not shown). BCA protein assay also showed thatSP1/fabric (w/w) ratio is approximately 2 mg protein/g fiber (2/1000).In parallel experiments it was demonstrated that L-1-SP1 (SEQ ID NO: 6)and L-4 SP1 (SEQ ID NO: 9) also bind to aramid (KEVLAR™). Followingincubation with L3-SP1/CNT aramid (KEVLAR™) fibers turned dark in color,indicating binding of the CNT thereto even after extensive wash.Incubation of 30 mg aramid with 180/1000 w/w L4-SP1-CNT dispersion,followed by bath sonication (90 min temperature ranging between 30-70°C.), fiber removal, extensive washing (using the buffer) and boiling (10min in 60 ul) to extract bound protein and CNT produced darkened fibersbearing bound protein as well as bound CNT.

CNT dispersion (0.1% CNT (Arkema, code C100), using L3SP1 (SEQ No 8))was incubated with aramid fabric (KEVLAR style 120 plain weave 195Denier, 58 g/m square; 22 ml suspension per g fabric) by agitation (1 h;25° C.; 150 rpm) followed by extensive wash in the same buffer, anddrying in the open air, overnight CNT content on fabric was about 9 mg/gfabric. Note that the bound CNT dramatically increases surface area, andthat the CNT are in close contact with one-another, affording improvedelectrical conductive properties.

Example 6 SP1 Variants Binding to Carbon Fabric

Carbon fabric is a well-known high-strength material with a variety ofimportant applications in aerospace and automotive fields, as well as insailboats and sport equipment, where its high strength-to-weight ratiois of importance. Continuous carbon fiber/epoxy composites have beenwidely used for structural applications due to their excellentmechanical properties. The polymer is most often epoxy, but otherpolymers, such as polyester, vinyl ester or nylon, are also used.However, their matrix-dominant properties, such as in-plane andinter-laminar shear properties, are much weaker than theirfiber-dominated properties, thus limiting the benefits of theseconventional composites. In addition, it is known that compositesexhibit lower longitudinal compressive strength, a matrix-dominatedproperty, than tensile strength.

CNT binding to the fabric via the protein increases its surface area,allowing better interaction with the fiber, and induces cross linkingbetween the fibers. In addition, protein binding to the fiber by itselfmay improve the interaction with the polymer through reactive groups onthe protein surface. It is demonstrated that some SP1 variants that bindCNT also bind to structural fibers.

Production of SP1-CBD Dissolved Inclusion Bodies Materials and Methods

SP1-CBD is expressed in bacterial hosts as insoluble inclusion bodies(IBs), as described in U.S. Pat. No. 7,253,341 to Wang et al. Briefly,SP1 cDNA encoding a 108 SP1 amino acid sequence (SEQ ID NO: 88) wascloned into an expression vector bearing a nucleotide sequence encodinga 163 amino acid CBD domain of Clostridium cellulovorans cellulosebinding protein A (SEQ ID NO: 87). The resulting nucleic acid constructencoded a SP1-CBD fusion protein which includes a peptide linker (SEQ IDNO: 89). Following cloning, the resulting plasmid was used to transformE. coli strain BL21 (DE3). Recombinant CBD-SP1 fusion protein synthesiswas induced in BL21 (DE3) by the addition of IPTG(isopropyl-D-thiogalactoside) to a final concentration of 1 mM tomid-log phase of the bacterial culture, followed by five additionalhours induction at 37° C. Recombinant SP1-CBD fusion protein (SEQ ID NO:86) was detected in inclusion bodies (IB), and the inclusion bodiesisolated and purified. Briefly, IBs containing SP1-CBD were dissolved inTrisma base (20 mM), NaOH (8 mM) (30, min on ice, 1:200 ratio (w/v)),followed by high speed centrifugation, 13,000 rpm for 30 min. Thesupernatent was diluted 1:10 in water and the pH was adjusted to pH=8.2(using NaPi buffer, 100 mM pH=6.8).

SP1 Polypeptide-CNT-Complex Binding to Carbon Fiber

Carbon (also glass and aramid) fabrics were washed with phosphate buffer(10 mM; pH 8) in a rotary shaking bath (160 r/min, 10 min, at roomtemperature) and then incubated in a rotary shaking bath (1 h each side,160 r/min, room temperature) containing aqueous SP1/CNT (SEQ ID NO: 8)suspension (suspension/fabric w/w ratio was 5:15; CNT concentration was0.05-0.4%, termed the “applied fraction”). The remaining suspension ofCNTs was termed the “unbound fraction”. SP1/CNT PAN fabrics were thenextensively washed with deionized water, termed the “wash fraction”,until the washing solution became colorless. The treated fabrics werethen air-dried for 1 week. To confirm and quantify CNT binding to thefabric, the amount of CNT in the “unbound

washed” fractions was subtracted from its amount in the “applied”fraction. For analytical purposes, the concentrations of SP1/CNT complexin dispersions were determined by two methods with similar results. Themaximal CNT binding as measured by this “subtraction” method was about 4mg CNT/gr fabric 0.4%:

-   -   1. Gravimetric method when the dispersion is coagulated &        flocculated by using HCl (0.3%), followed by centrifugation (10        min, 2500 g) and pellet dehydration in the oven (80 C,        overnight), followed by dry weight measurements.    -   2. Measuring absorption of diluted L3-SP1/CNT suspension before        and after its incubation with the fabric, including the fabric        washing solutions, using standard spectrophotometer or 96-well        plate reader at 405 nm    -   3. Bound SP1/CNT can be extracted from wet fabric using 0.5%        SDS, but not by using water. However, this extraction method is        not practical for evaluation of amount of bound SP1/CNT because        of the following reasons: even after very intensive extraction a        significant proportion of the CNT is still bound to the fibers        (evaluated by HR-SEM analysis); small broken pieces of fibers        are released by the extraction mixture.

TABLE 4 Characterization of dispersion made by conjugation of the SP1protein together with carbon nanoparticles (CNP; multiwall carbonnanotube and carbon black) Multi-walled CNT Graphi Carbon Black Carbonstrength ® Conductive CB CB CB nanoparticle C-100 Arkema N326 CB 220 550660 SP1 Variant Heat stable crude extract of recombinant L3SP1 proteinMaximal 2.9 ± 0.5 7 5.5 4.5 5.5 5.5 SP1/CNP dry weight/weight ratioDispersion Method Sonication¹ SP1/CNP  4%  17%  4%  4%  4%  4%concentration Fraction of CNP <25% <30% 45% 25% 20% 75% remain insupernatant upon centrifugation ² Long term <12 months <4 month ND ND NDND stability at room temp ³ pH stability 5.5-11 5.5-11 ND ND ND NDstability in the >0.3 mM >3 mM presence of divalent cation (Ca⁺² Mg⁺²)¹Sonication was conducted using a Hielscher Ultrasonics processorUIP1000 (1000 W), frequency: 20 kHz (autoscan), amplitude: 25 mm (85%),40 min sonication. ² Stability upon centrifugation, a quantify measurefor dispersion quality, was assessed by calculating the fraction of CNPremaining in the supernatant under centrifugation (dispersion wasdiluted to 0.1% in sodium phosphate buffer (10 mM; pH 8.0); Supernatantwas collected after centrifugation (30 min; 3000 g, in 50 ml Falcontube; 25° C.)). Both gravimetric and spectral methods were used todetermine the ratio of CNP concentration before and aftercentrifugation. Gravimetric method: CNPs were flocculated with 0.3% HCl,spun down (10 min, 4700 g), followed by pellet dehydrated (80 C.,overnight), and dry weight measurement (detonated supernatant (S)). Thesame procedure was conducted also on the dispersion beforecentrifugation (detonated total (T)). Stability upon centrifugation wascalculated according to the following equation: 1 − (T − S)/(T).Spectral method: optical density of diluted dispersion was determined at405 nm using 96 plate reader. Note that there is a linear correlationbetween CB concentration and optical density. Ratio of the opticaldensity of the dispersion before and after centrifugation wasdetermined. ³ Long term stability was determined upon long term storageof concentrated dispersion at room temperature as described in 2.

FIG. 1A is an HR SEM image of SP1/CB (N326) dispersion demonstratingthat the dispersion is nano-metric (particle/aggregate size is <500 nm).

FIG. 1B is an HR SEM image SP1/CB (N326) coating of a polyester fiber.

FIG. 2A depicts the effect of time of sonication on SP1/CB dispersion atdifferent SP1/CB concentration and dry SP1/CB w/w ratio.

The Spectro measurements relate to spectroscopic measurements and areindicative of CB concentration as a linear relationship of with opticaldensity. The optical density of diluted dispersion was determined at 405nm using a 96 plate reader. The ratio of the optical density of thedispersion was determined before centrifugation and several timesafterwards as a function of sonication as depicted.

Minimal time of sonication is 30 min at low dry SP1/CB w/w ratio (5.5)regardless SP1/CB concentration, but longer time of sonication (60 min)is required at higher dry SP1/CB w/w ratio (7). Maximal SP1/CNT andSP1/CB concentration are 4% and 17%, respectively. Long term stabilitytests at room temperature of 4% dispersion SP1/CNT and SP1/CB-N326 are<12 m and <4 m, respectively (Table 4). The SP1/CNT and SP1/CBdispersions are stable at wide range of pHs (pH=5.5-11) but precipitatein the presence of divalent cations (Mg+2 and Ca+2) in concentrationsof >0.3 mM and >3 mM, respectively.

SP1/CNP precipitation at low pH and in the presence of divalent cationshave several practical implications: for example, low pH and thepresence of divalent cations are useful conditions for CNP recycling andfor toxic waste reduction. All steps with the water based dispersion ofSP1/CNT and SP1/CB were carried out with de-ionized water (Concentrationof divalent cations <10 μM).

FIG. 2B is a plot depicting suspension as a function of sonication timefor various loading of CB in the SP1/CB complexes

Dispersion of Other Types of Carbon Black Conductive CB, CB220, CB550and CB660

The same methodology used for Graphistrength® C-100 and CB-N326, wasused to disperse other types of carbon black: conductive CB, CB220,CB550 and CB660. All CNPs were dispersed to some degree (Table 4). Thedifference in stability upon centrifugation may be attributed todifferences in specific gravity.

CNP/protein (Heat stable crude extract of recombinant L3SP1 protein)ratio is 2.8 and 7 for multiwall carbon nanotube (MWCNT) and carbonblack (CB), respectively. Maximal dispersion concentration is 4-5% and17% for multiwall carbon nanotube (MWCNT) and carbon black (CB),respectively. Note that for single wall CNTs which display much highersurface specific area, CNT/protein ratio is 8-fold lower. Therefore,much lower protein concentrations are needed in order to disperse CNPwith pure protein (MWCNT/protein ratio=20), but since proteinpurification is costly, crude extract is used without furtherpurification. In the absence of protein, the CNPs are not dispersed butrather readily precipitate on the bench. Dispersions stability for fewmonths, also at elevated temperature is essential for industrialapplication. The complexation of CNP with SP1 protein enables thedispersion of CNPs and enhances its stability. SP1/CNP complexprecipitate. Investigation of conductive CB will be further discussed.

Layer by Layer Coating

An important feature of the current invention is the “layer by layercoating” procedure, as described above, since it allows controlling CNPload on the yarn in a cost effective way minimizing waste, if any, ofexpensive nanoparticles. When low SP1/CNP concentration is applied(0.05% in dispersion; when liquor/fabric ratio=7 this value isequivalent to 3.5 g/Kg yarn) it is totally depleted from the appliedfraction and when PEI is used in the next step it allows binding ofhigher load of SP1/CNP (equivalent to 7 g/Kg yarn with total load of10.5 g/Kg), and so on. It is important to note that though increasingSP1/CNP concentration in the applied fraction may increase boundSP1/CNP.

Results

Kevlar treatment with both RFL and SP1/CNP don't change strength at maxload and elongation at break. SP1/CNP treatment don't change Strength at1% elongation (N)) (reduces flexibility), in contrast with the RFLtreatment that dramatically increase Strength at 1% elongation, which isimportant when the reinforcement cord is stretched.

Electrical Conductivity

FIG. 3A is a flow chart depicting a non-limiting general procedureemployed in the treatment of Polyethylene terephthalate (PET) yarn torender it electrically conductive in a range of 1 ohm/m to 20 Megaohm/m. In certain a certain embodiment resistance can range between 0.1Ω/m 10 mega Ω/m.

It should be appreciated that in certain embodiments, non-polymeric yarnis employed. As shown, the process is implemented in two primary stages;the SP1/CNP loading stage 15 and the copper loading stage 16, accordingto and embodiment.

SP1/CNP loading stage 15 stage commenced with the preparation of aSP1/CNP dispersion at step 20, as set forth above, to be loaded afterappropriate preparation of the PET yarn.

In step 21, the yarn was de-sized using soda ash triton or anotherscouring solution in an Ugolini dyeing machine, for example, with smartand controlled winding on bobbin to facilitate uniform coating.

In step 22, high molecular weight (60 KDa) branched PEI was dissolved incarbonate buffer (20 mM at a pH=8.8) and applied at 45° C. when thePEI/yarn ratio=0.035%-1.4% by weight.

In step 23, the yarn underwent extensive washing with carbonate bufferequilibrium to prevent the low concentration PEI to aggregate and toprecipitate CNP.

In step 24, the SP1/CNP dispersion is brought into contact with the yarnwhen CNP/yarn ratio=0.35%-0.9% by weight at 45° C. Typically thisprocess takes about 0.5-1 hour until complete depletion of the SP1/CNPfrom the dispersion is achieved.

In step 25 an evaluation of the loading of the composition coating isperformed. If the composition load is less than about 1 g of SP1/CNPcomposition/kg of substrate, for example, processing returns to step 22for additional PEI and CNP treatment. Depletion of the SP1/CNP isassessed by measuring the fraction of CNP remaining in the supernatantupon high-speed centrifugation under standard conditions (0.1% in NaPibuffer; 30 min at 3000 g, at room temperature), followed by flocculationof the supernatant with 0.3% HCl, centrifugation (10 min at 4700 g),pellet dehydration (80° C., overnight) and dry weight measurement.

If determined that that the SP1/CNP coating has achieved a thresholdthickness, operations continue to the copper loading stage 16. It shouldbe noted that various threshold thicknesses of SP1/CNP coating may beemployed in accordance with the particular application.

The copper loading stage 16 commences at step 26 with a PEI treatment tofacilitate bonding between the SP/CNP coating and palladium whenapplied. It should be noted that in certain embodiments other cationicpolymers are employed to facilitate bonding like copolymers and otherpolyamines.

In step 27, a Palladium coating is applied to the SP1/CNP layer bycontacting the coated SP1/CNP substrate with a Palladium solution.Palladium salt Sodium tetra-Chloro Palladate (NaPdCl₄) and PalladiumChloride (PdCl₂), sources were purchased from Acros and used asreceived. In one test run, NaPdCl₄ was dissolved in soft water inconcentration ranging between 0.01 mM-5 mM. All palladium solutionsprepared in PIPES buffer (20 mM, pH=6.8). In another test run PdCl₂ wasdissolved in water in presence of 5% technical ethanol without bufferaddition and used shortly after preparation.

In general, the concentration of NaPdCl₄ can range from 0.1 mM-1.0 mM.

In a first run, Pd application was performed in an Orbital ShakerIncubator machine at 45° C. and a speed of 140 rotations/hour in which1-10 meters of complex coated yarn was submerged in 100 mL palladiumsolution and shaken for five minutes.

In a second run, bulk experiments were performed using a Mini SymplexUgolini lab dyeing machine depicted in FIGS. 3B and 3C.

Specifically FIGS. 3B and 3C depict schematic views of a sample dyingmachine 30 processing during both in-out and out-in modes, respectively.Specifically, dying machine 30 is fitted with a perforated cylinder 32mounted in a treatment vessel 31 though which a process liquid 35 iscirculated by pump 33. Yarn 34 is wrapped around cylinder 32 duringprocessing and the treatment liquid 35 is pumped into cylinder 32through yarn 34 and the perforations in cylinder 32 when operating inout-in mode as depicted in FIG. 3B. In the in-out processing mode,liquid is pumped in the opposite direction out of cylinder 32 throughits perforations and through the yarn 34 as depicted in FIG. 3C. Itshould be appreciate that various dyeing machines providing suchfunctionality are may be employed.

As noted above, the bulk run was performed on 100 g of complex-coatedyarn submerged in 0.7 L Pd solution in a Mini Symplex Ugolini lab dyeingmachine with a kier volume of 0.7 L for about 0.5 hour at a liquor/yarnratio=7 with the following pump working parameters:

-   -   In-out time 2 min.    -   Out-in time 1 min.    -   Pump output 80%.    -   Pressure during the process ˜1 bar    -   Δ pressure ˜0.1 bar        The pH, the temperature, the pump output volume and pressure        were monitored in a data logging system. This step may be        repeated several times using water washes between the Pd        treatments for better uniformity.

Returning now to the process, in step 28, the Palladium coated substratewas reduced in a reduction bath. Reduction solution was prepared usingBorane dimethylamine complex (2.5 g/L), tri-Sodium citrate dehydrate (25g/L) and lactic acid (25 g/L). pH of 6.7 was achieved with addition ofNaOH (1M). All the reagents were purchased from Acros (Borane reagent),Merck (Citrate), Fisher Chemicals (Lactic acid) and used as received.The solution were stored for several days and found to be effective asit was prepared. Lower concentrations were also found to be effective aswell. It is possible to use even lower concentration of reductionsolution up to 10% of the standard solution.

It should be appreciated that in a certain embodiment, the applicationof a metallic catalyst, like Pd, can be achieved after a five secondscontact time of substrate and solution and in other embodiments theapplication can take as long as one hour.

In step 29, following a water wash, the Palladium coated substrate wassubject to a copper treatment though submersion in a copper solutionpurchased from Amza. The active solution includes the followingingredients:

-   -   A—(Sulfuric Acid Copper (2+) pentahydrate (10-25%),        Ethylene-dinitro-tetrapropan-2-ol (3-5%), methanol (1-2%))    -   B—(ethylene-dinitro-tetrapropan-2-ol (25-50%))    -   C—(Sodium hydroxide (25-50%))        Ingredients A, B and C were mixed before using in a standard        ratio: A (60 mL), B (60 mL) and C (25 mL) for about 1 L        solution.

The copper treatment was implemented in both bath and bulk processing.

Bath methods performed in 1 L glass Beaker containing 500 mL of coppersolution with mild stirring to prevent yarn damage. The glass bathheated directly on a heat plate to 40° C. and the temperature measuredinside the bath. The yarn was slowly dipped inside the bath whilestirring the Cu solution. The exemption of hydrogen gas exemplifiedreaction progress.

Generally, yarn remains in the Cu bath for a time period ranging from 30seconds to one hour depending on the target Cu thickness andconductivity. It should be appreciated that in a certain embodiment, theapplication of a metallic coating, like Cu, can be achieved after a fiveseconds contact time of substrate and solution and in other embodimentsthe application can take as long as one hour.

Following is summary of the relevant operational parameters employedduring the Pd coating, reduction, and Cu coating operation 37:

Step Bath Ugolini Machine Coating 1-10 m yarn, in 100 mL solution, 40°C. Gentle 100 g yarn (250-500 m), 0.7 L solution, conditions flow 140RPM, high Liquor/textile ratio = 700 37° C. Liquor/textile ratio = 7Pump work parameters: in-out time 2 min, out-in time 1 min, pump output80%, pressure during the process ~1 bar, Δ pressure ~0.1 bar. 38 0.01mM-5 mM NaPdCl₄ dissolved in PIPES 0.3 mM-1 mM NaPdCl₄ dissolved inPalladium buffer (pH 6.8, 20 mM), 5 min. PIPES buffer (pH 6.8, 20 mM),30 min. 39 5 mM-40 mM of Boron reagent concentration, 5 mM-10 mM ofBoron reagent, Lactic Reduction Lactic acid (8-80 mM) and tri SodiumCitrate salt acid (8 mM)and tri Sodium Citrate Bath (25-270 mM), pH 6.7was achieved by adding salt (25 mM), pH 6.7 was achieved by several mLof NaOH (1M). All the reagents were adding NaOH (1M). All the reagentsdissolved in soft water. 1 m yarn, 100 mL were dissolved in soft water.solution, 40° C. 140 RPM, Liquor/textile 30 min. ratio = 700, 1-5 min.Water 3 washes, 1 min each, with 100 mL water. 2 washes, 5 min each,with 0.7 L Wash water. 41 25° C.-75° C., 10 seconds - 40 minutes.Standard 45° C., 30 minutes. Standard Copper concentration (CuSO₄ × 5H₂O40 mM, ethylene- concentration. 1-4 repeats. Bath dinitro-tetrapropanol6 mM, NaOH 20 mM pH (CuSO₄ × 5H₂O 40 mM, ethylene- (CuSO₄) 13), 20% ofstandard. dinitro-tetrapropanol 6 mM, NaOH 20 mM pH 13 Final Water 3washes, 1 min each, with 100 mL water. 12 h 3 washes, 5 min each, with0.7 L Wash drying at 45° C. water. 12 h drying at 50° C.

-   -   Bulk processing was employed for the copper treatment coating        and it was found that three or four repetitions were required to        achieve the resistance of 2,000 Ω/m.

The copper coated yarn was dried in oven at 50° C. for 12 hours tostabilize it and X-Ray spectroscopy employed to measure the thickness ofcopper layer measured using Fischerscope™, after drying and generalmethod of thickness calculation “Cu/Base; plastic”.

An evaluation of the effects of CB and PEI depositions was performedusing 1100×2 dtex treated with 2 layers of CB 0.7 g/Kg and two layers ofand PEI of 0.14 g/Kg each. The concentration of PEI was 0.05%-0.0005%and the NaPdCl₄, reduction, and CuSO₄ baths were implemented inaccordance with procedures described above.

Effect of PEI, Amount of SP1/CB complex and CuSO₄ Bath Concentration onResistivity Effect of PEI at Constant Effect of SP1/CB Complex TimeSP1/CB Complex load Load at Two PEI loads in Cu 0.14% 0.07% 0.14% bathCB 0.35% 0.035% 0.0035% In process In process (min) PEI cap cap cap 0.7w % 0.35 w % 60 — 103 Uneven 30 13 182 Uneven 20 85 4600  92 4,60010 >20M >20M 290 >20M 7 — — 5 >20M >20M 755 >20M

The conclusion was that a concentration of 0.0035 g PEI/kg yarn was toolow and led to uneven coating for this particular trial; although, in acertain embodiment it such a low concentration of PEI is desirable. Theoptimal concentration for single CB layer was found to PEI be 0.0065g/kg yarn at the given experimental conditions. In a certain embodimentthe concentration ranges up to 10% w/w.

FIG. 4A is a chart depicting results from experimentation directed atestablishing optimal amounts of a competing agent CuCl₂ during Pdtreatment. The competing agent was employed to delay bonding of the Pdand the SP1/CNP complex until the Pd solution permeated through layersof yarn during treatment. Experimentation showed that Pd bonding in theabsence of competing agent resulted in a quick Pd bounding with theouter layers of yarn and insufficient bonding at the underlying layers.During experimentation, 1100×3 dtex polyester yarns were treated usingUgolini machine conducted in a bath for 10 minutes at 40° C. Theresistivity of the conductive yarns was then measured at various yarnlengths under tension of a 200 gram weight. Concentrations of NaPdCl₄and CuCl₂ reagents and corresponding resistance measurements anduniformity results are set forth in the following table:

NaPdCl₄ Treatment and Competing Agent With With No No CompetingCompeting Competing Competing Agent Agent Agent Agent 1 mM 1 mM 1 mM 0.3mM NaPdCl₄ + NaPdCl₄ + NaPdCl₄ NaPdCl4 10 mM CuCl₂ 3 mM CuCl₂ Resistance1.8 10⁶ 7.3 10⁷ 8.3 10⁶ 4.9 10⁷ Ω/m (AVG) Uniformity 118% 56% 17% 135%(CV)

As shown, the optimal ratio of reagents at these experimental conditionsis that producing the smallest CV of 17%

FIGS. 4A-4C are microscope images of 1100×3 dtex PET yarn enlarged 250×and depicts discontinuities in the copper coating. The orange-brownareas are characteristic of copper coating whereas the black areas arecharacteristic of carbon black underlayer.

The sample depicted in FIG. 4B was treated with 0.3 mM NaPdCl₄, whereasthe sample depicted in FIG. 4C was triple treated with 0.3 mM NaPdCl,and the sample depicted in FIG. 4D was treated with 1 mM NaPdCl₄together with 10 mM CuCl₂.

FIGS. 5A-5C depict the effect of temperature reduction from 45° C. to25° C. of the CuSO₄ bath treatment time when coupled with acorresponding increase in treatment time.

Specifically, FIG. 5A depicts an average resistance of 810Ω for a 10minute treatment time, whereas FIG. 5B depicts an average resistance of19Ω for a 30 minute treatment time, and 5C depicts an average resistanceof 13 for a 40 minute treatment. The conclusion being that lowertemperature of CuSO₄ bath (25° C. instead 45° C.) coupled with longertreatment time improves the uniformity up to Coefficient of Variation(CV) of 8%.

FIGS. 6A-6B are plots depicting current resistance for various PET yarnsegments as a function of tension. The tests were performed for coateddtex 1100×3 polyester yarn with high twist with the following coatings:

-   -   PEI (1^(st) and 2^(nd) layer 0.14 g/Kg, 3^(rd) layer 0.28 g/Kg,        layer 0.28 g/Kg)    -   Pd 1 mM (10 mole/Kg),    -   Reduction (5 mM of Boron reagent) followed with three short        washes    -   3 layers of CB (1^(st) layer 0.5 g/Kg, 2^(nd) layer 0.7 g/Kg,        3^(rd) layer 1.0 g/Kg) applied in an Ugolini dying machine.

Specifically, FIG. 6A depicts resistance response for yarns treated in aCu bath for only 10 minutes whereas FIG. 6B depicts the resistanceresponse for yarns treated for 40 minutes.

It was found that points of discontinuity of the copper coatingincreases electrical resistance. Tension applied on yarn compresses thefilaments together and improves contact between the fibers therebyimproving conductivity. Accordingly, it was found that increasingtension applied to high resistance yarn dramatically reduces resistance.However, conductivity of low resistance yarn did no significantlyimprove upon application of tension. Yarn with high level of twist alsodemonstrated an ability to reduce resistance emanating fromdiscontinuities in the copper coating for the reason noted above.

Fabrics

Several fabric types such as treated Polyethylene terephthalate (PET)fabrics and SP1/CNT treated Fiber Glass fabrics which were prepared inhouse involved to the

FIG. 7A is an image of three bobbins of PET yarn at various stages oftreatment. The left bobbin is a nude bobbin with neither SP1/CB nor Pdtreatments, whereas the middle bobbin has undergone full treatment; i.e.SP1/CB, Pd, and Cu treatments, and the right bobbin underwent SP1/CB andCu treatments without a Pd treatment. (All treatments were implementedwith an Ugolini dye machine.) It should be appreciated that SP1/CNTcoatings in place of SP1/CB will also provide the necessary pre-Pd andCu treatments.

Experimentation was performed to establish the necessity of each of theSP1/CB, PEI, and Pd treatments to achieved low resistance, uniformcopper coating when applied to a Cu bath. After KTS scouring and smartwinding of 1100×2 dtex polyester yarns, a first experiment lackingSP1/CB and Pd treatments, did not yield a Cu coating when a Cu bath wasapplied to the yarn. A second experiment performed with SP1/CB treatedyarns; but, without Pd and Redox layers, also did not yield a Cu coatingwhen a Cu bath was applied. A third experiment in which the yarn wassubject both SP1/CB and Pd treatments yielded suitable for Cu coatingwhen treated with a Cu bath.

Following is test data indicating the necessity of SP1/CNP and PEI, andPd treatments as prerequisite treatments for a successive Cu treatmentproviding a uniform low resistant conductive coating.

PET 1100x2 PET 1100x2 PET 1100x2 dtex, dtex, dtex, treated treated withuntreated with SP1/ SP1/CB + PEI and Yarn type yarn CB + PEI NaPdCl₄ CBand PEI treatment No 2 layers of 2 layers of CB, PEI CB, PEI NaPdCl₄ andreduction No No 1 mM NaPdCl₄ - treatment in Ugolini 30 min, reduction -30 min CuSO₄ treatment in 30 min, 3 30 min, 30 min, 3 repeats Ugolinirepeats 3 repeats Ω/m R > 2 * 10⁸ R > 2 * 10⁸ R = 10²-10³ Resistance

In conclusion, an effective Cu coating is achievable only after SP1/CB,PEI, NaPdCl₄, treatments. In certain embodiments reduction treatment isalso applied after the Pd treatment whereas in other embodiments thisreduction treatment is not employed. The absence of any of thesetreatments will prevent the application of a low resistance, uniformcopper coating.

FIGS. 7B-7E are enlarged views of the fibers of the properly Cu coatedyarn the middle bobbin of FIG. 7. Specifically, the 226 dtex yarndepicted in FIG. 7B exhibited a resistance of 150 Ω/m after a fiveminute Cu bath, whereas the 1100×2 dtex yarn depicted in FIG. 7Cexhibited a resistance of R=400 Ω/m after a one minute Cu bath, and the1100×2 dtex yarn of FIG. 7D exhibited a resistance of R=10 Ω/m after afive minute Cu bath.

FIG. 7E is an enlarged view of a 1100×2 dtex yarn coated in CuSO₄ bathyarn in having a resistance of 40 Ω/m achieved with a Cu coatingthickness of 3.5 in some areas and 3.2 micron in other areas whenmeasured by X-RAY XDL-B instrument “Fischerscope”. Dark areas have ameasured thickness of 1.78 micron.

FIG. 8 is a plot depicting resistance/length of KEVLAR™ yarns as afunction of treatment time in a Cu bath. This experimentation wasperformed using 1100×4 dtex yarns with 3 layers of SP1/CB complex andPEI prior to treatment in an CU bath as described above. KEVLAR™ yarnsexhibited high conductivity. Measurements of resistance were performedwithout the addition of weight.

Similar experimentation was performed for polyester fabrics.Conductivity was found to be strongly depended on NaPdCl4 concentration,CuSO₄ treatment time and addition of a competing agent.

FIG. 9A is a plot depicting resistance/length of conductive polyesterfabrics as a function of treatment time in a NaPdCl₄ bath. Surprisingly,it was found that decreasing the NaPdCl₄ concentration from 1 mM to 2 μMincreased the resistance as depicted.

FIG. 9B is a picture of several fabric samples after various stages oftreatment from samples 1 to sample 4 having a complete copper coating.

Experimentation was also performed for glass fiber fabrics in which twolayers of SP1/CNT were deposited on the glass fabric, treated with PEIfollowed by a NaPdCl₄ bath following by reduction and washes. Cu coatingperformed in bath at different time periods ranging from 3 minutes to 10minutes. Additionally the experimentation of fabrics without the PEItreatment were also performed.

FIG. 10A is a plot depicting resistance/length of conductive glass fiberfabric as a function of treatment time in a CuSO₄ bath.

FIGS. 10B and 10C are images of samples after different treatment stagesfor operations including PEI treatment and without PEI treatment;respectively. As noted above, the presence of the rust or copper coloris characteristic of the presence of the copper coating. As shown in thecontrast in color of sample of each of the experiments, the lighter rustcolor in FIGS. 10B and 10C is characteristic of a more complete andcopper deposition.

Conductive Carbon Black (CB_(max))

As noted above, dispersion of conductive carbon black (CB_(max)) wasinvestigated. It was found that PET yarn loaded with a nanometricdispersion of SP1/CB_(max) complex yielded uniform and conductivecoating even in the absence of electroless copper electroless coating.

Furthermore, it was found that copper electroless deposition on yarnloaded with SP1/CB_(max) complex facilitated higher coating uniformityof copper coating reduced resistance than that of yarn loaded withnon-conductive CB.

During investigation, a nanometric dispersion of conductive SP1/CB_(max)complex was produced from CB_(max) obtained from Cabot Corporation(available at 2 Seaport Ln #1300, Boston, Mass.) through the proceduredescribed above.

Two parameters were tested; the quantity of SP1/CB_(max) deposition andthe thickness of the yarn. Three grades of yarn were investigated; thickyarn having a dtex of 1100×3, medium yarn having a 1100×2 dtex, and thinyarn having a dtex of 1100×1.

Each yarn grade was treated with several layers of SP1/CB_(max)dispersion. The final results are summarized in the below table.

Resistance for SP1/CB_(max) Complex Without Copper Coating 1 CB_(max)layer 2 CB_(max) layers 3 CB_(max) layers dtex R (MΩ/m) % CV R (MΩ/m) %CV R (MΩ/m) % CV 1100x1 >200 — >200 — 181  26% 1100x2 99 7.3% 21.7 10.1%16.5 8.8% 1100x3 — — 10.6 11.6% 9.5 7.0%

Initial results indicate that the coating levels of thin yarns (1100×1)were too low as may be seen by the high resistance. However medium andthick yarns (1100×2 and 1100×3) showed significantly lower resistanceand a very low % CV. This is especially significant when contrasted withthe high resistance (R>2*10⁸ Ohm/m) measured in yarns loaded withnonconductive CB/SP complex.

The added coating uniformity and resulting resistivity reductionachieved through SP1/CB_(max) complexes is crucial in the achievement ofhigh conductivity using copper electroless coating yarns and fabrics.

Additionally, various yarn grades loaded with SP1/CB_(max) complex wereelectrolessly copper coated.

Standard copper solutions were obtained from Amza Ltd. (Available at 37Nachshon St. Industrial Area Sgula, Petach Tikva, Israel).

The active solution included three ingredients; sulfuric acid copper(2+) pentahydrate (10-25%), ethylene-dinitro-tetrapropan-2-ol (3-5%),methanol (1-2%), ethylene-dinitro-tetrapropan-2-ol (25-50%), and sodiumhydroxide (25-50%). These ingredients were mixed before using in astandard ratio: A (60 mL), B (60 mL) and C (25 mL) for 1 L solution.

Multiple short coating steps of about ten minutes each with dilutedcopper solution prevented material waste and also facilitated uniformcoating.

The copper was deposited on yarns coated with nonconductive SP1/CBcomplex using a batch dying machine as described above and it was foundthat yarn crossover occurring when wound around the bobbin preventedcompletely uniform coatings.

Surprisingly, the SP1/CB_(max) complex facilitated a higher coppercoating uniformity; thereby further reducing resistance, even in thepresence of yarn crossover.

The resulting resistance measurements and associated coefficient ofvariation are set forth in the table below.

Resistance for SP1/CB_(max) Complex With Copper Coating Before Cutreatment After 3 layers of Cu Uniformity Uniformity dtex CB layers R(MΩ/m) % CV R (MΩ/m) % CV 1100x2 3 16.5 8.8% 1.2 11.1% 1100x3 2 10.611.6% 3.6 12.7%

As shown in this table, SP1/CB_(max) complex is a highly effectiveplatform for achieving uniformly conductive yarn or fabrics even afterapplication of metallic coatings.

Polymeric Protection

Low resistance yarn or fabric of about 10-1,000 Ω/m is achieved throughthough the depositing of several layers of copper through 10-18treatments of diluted copper solution, according to a certainembodiment. The yarns and fabrics are susceptible to detachment of thecopper coating from the fiber through mechanical abrasion.

Accordingly, polymeric coating, like polyurethane for example, wasapplied after copper deposition and washing. As expected, resistance wasincreased; however, the benefit of the added durability offset theincrease in resistivity.

It should be appreciated that in a certain embodiment FeCl₃ is used aconductive agent. Advantages of us using FeCl3 as a conductive agent isthat it is more inexpensive than Pd and Cu, only one treatment isrequired, a variety of polymers are suitable for FeCl3 attraction, andalso leads to leads to lower conductivity as required in certainapplications.

In another embodiment, aluminum is employed with suitable polymericsubstrates like polypyrrole or polyaniline.

In another embodiment, nickel with suitable polymeric substrates likepolypyrrole or polyaniline.

In another embodiment, cobalt with suitable polymeric substrates likepolypyrrole or polyaniline.

In another embodiment, gold metals with suitable polymeric substrateslike polypyrrole or polyaniline.

In a certain embodiment the carbon particles are implemented asmicro-particles instead of nanoparticles.

Although the invention has been described in conjunction with specificembodiments thereof, it is evident that many alternatives, modificationsand variations will be apparent to those skilled in the art.

In certain embodiments, the outer coating is constructed from silver,other embodiments employ platinum and other embodiments employ Rhodiumor Iridium. In certain embodiments a different metals are employed inseparate coatings whereas in certain other embodiments identical metalsare implemented as separate coatings. In other embodiments the coatingsare implemented as metal alloys.

In some embodiments the thickness of the outer metal coating isimplemented between 0.1 μm and 10.0 μm whereas in other embodiments thethickness of between 0.01 μm and 100.0 μm.

A certain embodiment uses a twist level of either fibers or yarn as lowas 20 twists/meter whereas different embodiments use twist levels ofover 100 twist/meter. It should be appreciated that other twist levelsare included in the scope of the invention.

Accordingly, it is intended to embrace all such alternatives,modifications and variations that fall within the spirit and broad scopeof the appended claims.

All publications, patents and patent applications mentioned in thisspecification are herein incorporated in their entirety by referenceinto the specification, to the same extent as if each individualpublication, patent or patent application was specifically andindividually indicated to be incorporated herein by reference. Inaddition, citation or identification of any reference in thisapplication shall not be construed as an admission that such referenceis available as prior art to the present invention. To the extent thatsection headings are used, they should not be construed as necessarilylimiting.

While certain features of the invention have been illustrated anddescribed herein, many modifications, substitutions, changes, andequivalents will now occur to those of ordinary skill in the art. It is,therefore, to be understood that the appended claims are intended tocover all such modifications and changes as fall within the true spiritof the invention.

What is claimed is:
 1. A conductive yarn comprising: a plurality ofinterlocked fibers at least partially coated with a composition ofcarbon black (CB) and SP1 variant (SP1/CB); one or more polyaminecoating(s); and an outer metal coating.
 2. The conductive yarn of claim1, wherein the polyamine coatings are a first polyamine coatingsandwiched between the fiber and the composition and a second polyaminecoating between the composition and the outer metal coating.
 3. Theconductive yarn of claim 1, further comprising an inner metal coatingdisposed between the second polyamine coating and the outer metalcoating.
 4. The conductive yarn of claim 1, wherein said SP1 variant isnon-covalently bound to the carbon black.
 5. The conductive yarn ofclaim 1, wherein said SP1 variant has the amino acid sequence of SEQ IDNOs: 3, 4, 6, 8, 9, 14-18 or 86; wherein the carbon black is selectedfrom the group consisting of conductive carbon black and non-conductivecarbon black; wherein the outer metal coating is a copper coating;wherein the inner metal coating is Pd(II), Pt(II), Rh(I), Ir(I), iron,aluminum, gold, silver, nickel, or combination thereof; wherein thefiber is selected from the group consisting of cotton fiber, wool fiber,silk fiber, glass fiber, nylon fiber, polyester fiber, aramid fiber,polyethylene fiber, poly-olefin fiber, polypropylene fiber, and elastanefiber; wherein the load of said SP1/CB on said yarn is between 0.01gr/kg and 100 gr/kg; wherein said CB:SP1 ratio is between 0.1:1 to 30:1dry w/w; wherein the yarn has a twist range of at least 10 winds/meter;wherein the outer metal coating has a thickness of between 0.01 μm and100.0 μm; wherein said yarn has a resistance between 0.001 Ω/m to 1000mega Ω/m; wherein said polyamine includes polyethyleneimine (PEI);wherein the yarn further comprises a polymeric coating on the outermetal coating; or any combination thereof.
 6. The conductive yarn ofclaim 5, wherein the load of said SP1/CB on said yarn is between 5 gr/kgand 15 gr/kg; said CB:SP1 ratio is between 2.5:1 to 8:1 dry w/w; orcombination thereof.
 7. The conductive yarn of claim 1, wherein theinner metal coating is Pd(II), Pt(II), Rh(I), Ir(I), iron, aluminum,gold, silver, nickel, or combination thereof.
 8. A conductive yarncomprising: a plurality of interlocked fibers at least partially coatedwith a composition of conductive carbon black (CB_(max)) and SP1 variant(SP1/CB_(max)); one or more polyamine coating(s); and an outer metalcoating.
 9. The conductive yarn of claim 8, further comprising apolymeric coating on the outer metal coating; wherein the polyaminecoatings are implemented as a first polyamine coating sandwiched betweenthe fiber and the composition of the SP1/CB_(max) and a second polyaminecoating between the composition and the outer metal coating; orcombination thereof.
 10. A conductive film comprising: a polymeric filmcoated at least partially coated with a composition of carbon black (CB)and SP1 variant (SP1/CB); a plurality of polyamine coatings; and anouter metal coating disposed on the polyamines coating.
 11. Theconductive film of claim 10, wherein the polyamine coatings are a firstpolyamine coating sandwiched between the film and the composition and asecond polyamine coating between the composition and the outer metalcoating.
 12. The conductive film of claim 10, wherein the inner metalcoating is disposed between the second polyamine coating and the outermetal coating.
 13. The conductive film of claim 10, wherein the CBincludes conductive CB_(max).
 14. A method of producing the conductiveyarn of claim 1, comprising: contacting a plurality of fibers with apolyamine so as to form a plurality of polyamine coated fibers;contacting the polyamine coated fibers with a dispersion comprising anSP1/CB complex so as to form a plurality of complex coated fibers; andcontacting the plurality of complex coated fibers with a metalliccatalyst so as to form a metallic catalyst coating on the plurality offibers.
 15. The method of claim 14, further comprising at least one stepof washing said yarn with a buffer or water after each step of themethod; wherein the contacting the plurality of complex coated fiberswith a metallic catalyst is implemented in a textile dying machine or abath; wherein said dispersion comprises an SP1/CB composition in aconcentration of between 0.01% and 50% w/w; wherein said dispersioncomprises an SP1/CB composition is in a concentration of 2%, 4%, 6%, or8% w/w; wherein said PEI is applied to the yarn at a load of 0.0035% to30.0% w/w; wherein said method is performed at temperature between4°-90° C.; wherein the metallic catalyst is iron, gold, silver, nickel,platinum, aluminum, or any combination thereof; wherein the metalliccatalyst is implemented as Pd(II), Pt(II), Rh(I), Ir(I), Cu(II) or anycombination thereof; wherein the method further comprises applying anouter metallic coating of copper outside of the metallic catalystcoating the fibers; wherein the method further comprises drying thefibers after contacting the metal catalyst; wherein the CB of the SP1/CBcomplex is implemented as conductive carbon black (CB_(max)); or anycombination thereof.
 16. The method of claim 15, wherein said dispersioncomprises an SP1/CB composition is in a concentration of 0.05%, 0.1%,between 0.05% and 0.15%, or between 1% and 20% w/w; wherein said PEI isapplied to the yarn at a load of 0.007% to 0.7% w/w; wherein themetallic catalyst is a combination of Pd(II) and Cu(II); wherein thecontacting the plurality of fibers with a metallic catalyst isimplemented in a time span ranging from 5 seconds to 1 hour; wherein themethod further comprises reducing the metallic catalyst to the metallicstate; wherein the method further comprises applying a polymeric coatingonto the outer metallic coating of copper; or any combination thereof.17. The method of claim 16, wherein the ratio between Pd(II) and Cu(II)is 1:10.